Corporations’ activities are a substantial cause of environmental damage and associated human suffering. However, conventional management thought assumes that as companies become more environmentally-friendly, profits diminish. The impacts of climate change have become increasingly apparent, and many are asking what can be done to ensure the quality of life of future generations. If humanity is going to slow climate change, corporations will have to play a major role. Our research goal is to investigate if there is a statistically significant correlation between corporate environmental performance and financial performance. Many environmental-financial performance researchers have measured environmental performance using managerial opinion surveys, but a respondent’s opinion can differ from a firm’s actual performance. We have collected environmental performance metrics, which decrease measurement bias by reducing the judgment involved with measurement. Lastly, many quantitative studies have not examined the environmental-financial performance relationship over time, while our study uses data from 2008 – 2013. Our research team will test the following hypotheses: a) higher corporate environmental performance is associated with higher financial performance; b) higher corporate environmental performance is associated with lower financial performance; and c) there is no relationship between corporate environmental performance and financial performance. To test these hypotheses, we have collected greenhouse gas emissions, energy use, revenue, and net income data from corporate sustainability and financial reports. We will use SPSS to analyze the relationships between environmental and financial variables with correlation statistics and regression. We anticipate positive correlations between the environmental and financial variables. If we discover positive correlations, then it will suggest that a profit incentive accompanies reduced energy use and emissions. Should corporations buy into this incentive, it could lessen the need for policies like carbon taxes that incentivize firms to reduce emissions.

Around 2 million people receive physical therapy (PT) every day and this number continues to increase. Yet current PT practices often leverage low-tech guidance, involve minimal patient-therapist communication and do not provide means for accountability. Thus the problem we seek to solve is how to modernize the PT experience to enhance the relationship between therapists & patients and equip patients to better manage their therapy routines. In response, Achilles is a physical therapy prescription system leveraging professional expertise to provide on-demand guidance throughout the rehabilitation process. This is accomplished via software that crowd-sources physical therapy demonstrations to provide a library of guided exercises for physicians to ‘prescribe’ to their patients. As a result, physicians can see when patients are doing their exercises, creating accountability. Additionally, the final design gamifies the physician and patient experiences, encouraging a regular PT routine and increased communication on both ends. Achilles modernizes the physical therapy experience and in turn, patients are able to better manage their therapy, recover faster and avoid recurrence in the future.

Many youth in residential treatment engage in non-suicidal self-injury (NSSI), a deliberate act of self-harm without the intent to die, which holds serious and potentially lethal consequences. Prevalence rates of residentially-treated youth engaging in NSSI have been on the rise and it is increasingly becoming a topic of concern among practitioners. Understanding the risk factors for NSSI is an important first step in preventing this kind of behavior. The study examined the relationship between the frequency, severity and method of NSSI against age, gender, ethnicity, and history of mental health diagnoses by performing data analysis on 112 discharged clients gathered from a residential treatment facility in the Pacific Northwest. This study found 38% of clients engaged in NSSI and a number of correlations were additionally examined. The study allows for an exploratory look into NSSI within residential treatment, suggests an earlier age of onset for this behavior, and additionally reveals previously unrecognized NSSI methods that hold future implications for practitioners. Understanding the frequency and types of youth most vulnerable to engaging in NSSI can assist in establishing a foundation for practitioners to provide more effective services and reduce the behavior overall.

Unlike differentiated germline cells, germline stem cells (GSCs) do not undergo apoptosis when treated with chemotherapeutic agents and gamma-irradiation. Similarly, cancer stem cells are thought to be particularly resistant to apoptosis, a phenomenon that can result in tumor relapse. Our research aims to identify the molecular mechanisms that allow stem cells to survive these stressors and re-populate the tissue. To this end, we screened a panel of 500 small molecules, provided by the National Cancer Institute, to identify agents that can induce apoptosis in GSCs. To this date, we identified two possible compounds that induce death caspase-1 activation and therefore apoptosis in GSCs. We are currently working to identify the pathways targeted by these molecules to understand how GSCs avoid apoptosis. Our second goal was to examine the mechanisms that allow stem cells to exit from their quiescent state and repopulate the germline tissue. Specifically, we examined autophagy as one candidate for having a role in the regeneration process. To do this, we compared the regeneration rates of wild-type (WT) and Thor2-/- mutant flies, which exhibit reduced levels of autophagy. We administered 50 Gy of γ-irradiation to female WT and Thor2-/- mutants to damage the germline tissue. We then sacrificed flies after 0, 2, 3, 5 and 7 days to examine the level of regeneration in the tissue. Our findings suggest that Thor2-/- mutants repair the germline tissue more slowly than WT flies, indicating that autophagy has a role in the regenerative process. In the future, we plan to repeat this experiment using Atg1 and UVRAG mutants to confirm that autophagy is required for an efficient and rapid regenerative response. In addition, we will conduct a genome-wide screen to identify other processes that control the transition from quiescence to rapid division during the regenerative process.

Landscape architecture has historically concentrated on designing aesthetically pleasing gardens, boulevards, and parks, which has resulted in a clear separation between how nature is perceived and used in urban environments. Only recently have designers stressed the importance of creating urban landscapes that mimic natural systems to provide aesthetic, social, and ecological function. Apart from aesthetics, designers try to create dynamic landscapes that can help mitigate the effects of urban runoff, climate change, and habitat degradation from humans. Keystone species generate habitat and influence ecological processes in natural environments. Beavers are both keystone species and ecological engineers that create complex wetland habitat types, which improve spawning for fish, increase vegetation in riparian zones, and trap sediment to increase nutrients within the stream. Once prevalent in today’s urban areas, beavers have often been in conflict with humans by causing flooding, damaging infrastructure, and cut down trees. However, even in an urban environment, beavers can provide ecosystem benefits by slowing urban runoff, raising the water table, increasing water filtration and retention, and mitigating the effects of climate change. Beavers can be integrated into the design of new and established urban green spaces to improve ecosystem processes. In this study, we examine how the field of landscape architecture is approaching the use of beavers in the design of wetlands and urban natural areas at regional and site levels. We present a number of case studies that identify various approaches, successes, and failures in incorporating beavers into project designs.

Past studies have shown that plasma created in air can effectively be used to modify the velocity and shape of atmospheric shock waves.The aim of the research is to assess the effect of plasma density distribution on the shock front development, or more specifically to study the effect exponentially increasing density within the sphere will have on the shape and velocity of the wave. For this, the problem was first solved analytically and then numerical simulations were performed to demonstrate the effect in a range of shock velocities and varying conditions within the plasma region. The results of these findings can find potential use in Astrophysics for shock waves generated within the stellar interior, Magnetohydrodynamics as well as for aerodynamic flow control in trans- and supersonic flights by creating plasma around a moving vehicle.

Tissue engineering has a large potential in the medical field because it can utilize the growth of stem cells to treat illnesses related to organ failure. However, researchers still lack the tools necessary to control the dynamic presentation of signals presented to a stem cell in its microenvironment. The DeForest lab is tailored towards addressing the issue of controlling cell growth in 4 dimensions—time and 3D space—by using a combination of modified proteins and light-programmable hydrogels. By combining the two, we hope to be able to control the cellular microenvironment in order to direct stem cell behavior. This can be accomplished by using protein constructs modified with a bioorthogonal reactive handle and photopatterning. My particular research involves creating, expressing, and purifying modified protein constructs from the DNA sequences for mCerulean (a blue fluorescent protein) and mCitrine (a yellow fluorescent protein), which represent model growth factors to direct stem cell fate. Towards this, I performed standard molecular biology techniques such as PCR, agarose gel electrophoresis, digestion, ligation, and transformations. In performing transformations with competent cells, those with porous membranes, we expect to have cells that have taken in the plasmid and thus can express the desired fluorescent proteins. Through an enzymatic reaction, we are able to introduce a bioorthogonal synthetic handle site-specifically onto our proteins. Photopatterning is performed by including a photocaged reactive group within the hydrogels, which upon UV light exposure will uncage the reactive group allowing for a reaction to occur with the synthetic handle. If this is successful, we plan to continue with growth factors TGFβ and BMP2. This research will have a massive impact in tissue engineering because it will allow for the recreation of physiological structures, grown outside the body with the patient’s own cells, and then implantation of the structure to the patient’s needs.

Hypoxemia, abnormal heart rate, and high capillary refill time are all signs of poor health. There is currently no single device that can evaluate all three key vital signs. We developed algorithms and explored the potential to measure these signs with conventional pulse oximetry finger clips consisting of two LEDs on one side (red and infrared) and a light sensor on the other side. We took a retired pulse oximetry finger clip, fit it with our hardware, and used an Arduino microcontroller and MATLAB software on a computer to process the data via a serial connection. The ratio of red to infrared light transmitted through the pulsatile blood was used to calculate the concentration of hemoglobin in the arteries, following modern pulse oximetry methods. The heart rate was analyzed with our custom spatial and frequency domain signal processing. The capillary refill time was measured by pressing and releasing the finger, and measuring the time it took for the transmitted light to return to pre-pressure levels. The system was able to resolve blood pulsatile changes in the red and infrared wavelengths. Initial testing on healthy controls revealed reasonable accuracy for pulse oximetry, high accuracy and precision for pulse rate, and high fidelity for the novel measurement of capillary refill time. Additional work is necessary for calibrating the device for pulse oximetry measurements. Likewise, further testing is required to fully characterize system performance. We constructed a prototype device and demonstrated initial feasibility for measurement of blood oxygenation, heart rate, and refill time. Because of the low cost of the materials involved and the improved accuracy of the sensor, this device could be a useful tool for hospitals in diverse locations.

Social scientists have a longstanding interest in the birth and death of social movements. Recently, some scholars are re-imagining these theories to accommodate the growing adoption and use of Internet-based communication technologies such as social media, hypothesizing about the ability of these technologies to shape and sustain movements. YesAllWomen is one example of a protest that took advantage of the widespread and distributed characteristics of dialogue offered on Twitter. This conversation, which grew out of a direct response to offline events, reached large audiences in a short amount of time. In current theories of movement success, leadership is a necessity. However, many have argued that these theories are out of date and must be extended to account for changing communication frameworks. This study examines #YesAllWomen in its beginning stages to understand the formation of leadership structures and Twitter’s role in the movement. The study takes a mixed method approach. Using a random sample of tweets from the first two days following the creation of the #YesAllWomen hashtag, the study identifies important individuals and explores how they represent leaders with or without taking a hierarchal role. Quantitatively, this study utilizes social network analysis to identify important positions within the conversational network. The two methods work together to show what the structure of leadership looks like in this context and who the individuals are who play a central role in the movement over time. Ultimately this study provides a unique view into a modern social movement, and seeks to identity shortcomings in social movement theory.

Vascular calcification greatly increases risk of cardiovascular disease, the leading cause of death in patients with diabetes and chronic kidney disease, by decreasing the compliance of major arteries. Vascular calcification is an active process attributed to vascular smooth muscle cell (VSMC) mineralization. The type III sodium-dependent phosphate cotransporter, PiT-1, regulates osteochondrogenic conversion of VSMCs under elevated extracellular phosphate conditions. Studies in our lab found that VSMCs respond to elevated phosphate through PiT-1 with a cell signaling mechanism independent of phosphate uptake. However, the signaling pathway by which PiT-1 induces osteogenic gene expression in VSMCs is poorly understood. I hypothesize that the intracellular domain of PiT-1 is required to bind adaptor proteins in response to elevated phosphate, triggering a phosphorylation cascade that promotes osteochondrogenic differentiation. In this study, VSMCs were engineered to overexpress the intracellular domain of PiT-1 in the cytosol to test the hypothesis that this domain would bind to adaptor proteins and inhibit VSMC calcification. Primary murine aortic VSMCs were transduced with the PiT-1 intracellular domain by retroviral infection. RNA overexpression of the intracellular domain was confirmed by reverse transcriptase PCR. However, expression of the protein measured by Western Blot was much lower than expected. The VSMC strains were induced to calcify at 3.0 mM extracellular phosphate. VSMC strains had significant induction of calcification at 3.0 mM elevated phosphate (VC = 90.09 ± 9.29, inPiT-1 = 71.89 ± 11.97 ug Calcium/mg Protein) compared to 1.0 mM phosphate, but calcification was not significantly different between VSMC strains. These results suggest that the amount of expression of the intracellular domain construct in the engineered VSMCs was not large enough to determine the effect of this protein on vascular calcification.

Gas chromatography (GC) is a powerful analytical tool that can be used in a wide range of fields to analyze a myriad of different volatile compounds. However, it generally requires 10-60 minutes to obtain a suitable chromatogram using conventional GC, which can be impractical if a large number of samples are to be analyzed. Using high speed gas chromatography, specifically Low Thermal Mass Gas Chromatography (LTM-GC) with valve-based injection, chromatograms with suitable information content can be obtained in a fraction of the time required using conventional GC. With LTM-GC, chromatographic separations that may take 10-60 min using conventional GC may only require 0.5-2 min. By obtaining and critically examining the data from chromatograms of standard mixtures and essential oils using both conventional GC and high speed LTM-GC, this study demonstrates the advantages and potential for the LTM-GC technology for enabling high throughput experimentation.

Previous work to investigate grassland evolution in Earth’s earliest grasslands, in southern Argentina, has yielded competing ideas about how changes in vegetation were correlated with climate changes, especially moisture availability. Among the proxies previously investigated, data on the abundance of phytoliths (microscopic biosilica bodies found in cells and tissues of vascular plants) were collected from sections of sediment deposited between about 20 to 40 million years ago in the Gran Barranca region of southern Argentina. Phytolith abundances, particularly of grasses and palms, were compared with indicators of past aridity, including sedimentological and isotopic data. Different metrics have thus far yielded differing interpretations, and the relationship between the abundance of certain plant taxa and moisture availability remains unclear. Diatoms are also present in the sediment sections, although it is not a lacustrine sequence. Among the diatoms present in the section, those of the genus Orthoseira, known to prefer wetland/bog habitats may be among the most useful. Other common genera include Staurosira, Navicula, and other fairly ubiquitous benthic taxa as well as the planktonic diatom Aulacosiera in some sections. Here we compare the abundance of palm phytoliths with the abundance Orthoseira, as well as the ratio of benthic to planktonic diatoms. If high planktonic to benthic ratios and/or high prevalence of Orthoseira co-occur with times of inferred transition to palm forest, the diatoms may lend a line of evidence to suggest that humid, wet conditions occurred with the spread of palm forest.

Southern Resident killer whales (Orcinus orca) occur seasonally in the inland waters of Washington, which consist of the Strait of Juan de Fuca, the San Juan Islands, Georgia Strait, and Puget Sound. They are most frequently present in the inland waters in the summer months, in part because their movements and core habitat are influenced by the availability and distribution of Chinook salmon (Oncorynchus tshawytscha). Southern Resident killer whales were listed as endangered under the federal Endangered Species Act in 2005. Recent downturns in population abundance of Southern Residents, from 87 whales in 2010 to 78 whales at the end of 2014, have highlighted the need to better understand seasonal distributions of this threatened population. Specifically, I am interested in quantifying how the occurrence of Southern Residents in inland waters has changed over time and whether there has been a shift in seasonality. My research focuses on the sub-pod known as J pod because it is most often observed in inland waters year-round. The Whale Museum in Friday Harbor, Washington compiled data for reported killer whale sightings for the years 1976-2013. Using the statistical software package R, I used generalized linear models to model the relationship between month, year, and sightings to determine how month and year affected the number of J pod sightings and the seasonal presence of J pod in inland waters. Initial results show that there are seasonal trends in sightings from year to year. Studying patterns in Southern Residents’ seasonal presence in inland waters will allow for the formulation of hypotheses for reasons why seasonality has changed, such as changes in the condition and timing of salmon runs. Thus, my results will provide insight into various factors affecting Southern Resident distribution, allowing for more targeted protection of the population.

The HIT (Helicity Injected Torus) research lab at UW has been remarkably successful in demonstrating a novel concept for fusion plasma confinement, which if proven, would be considerably cheaper than any other fusion concept currently being researched. However, validating theoretical and predictive claims is dependent on our ability to accurately characterize the experiment. The IDS ( Ion Doppler Spectrometer ) diagnostic provides this ability, recording information on ion flow and temperature. This presentation will outline results from IDS on the HIT group’s current and previous experiments (HIT-SI and HIT-SI3) with comparison to numerical simulations, and results of the comparison tests between Coherent-IDS ( C-IDS ) and MultiChord-IDS (MC-IDS) for potential use on HIT-SI3. Data from HIT-SI found ion temperatures in the range of ~10-20eV, and dynamic ion velocities of ~10km/s. Comparisons with NIMROD and PSI-TET simulation codes found strong agreement in relative temperatures and velocities, across the geometry of the experiment, but lack of agreement in magnitudes, motivating the addition of a neutral fluid to the simulated plasma. Data from HIT-SI3 shows temperatures of ~10-15eV, and velocities of ~3km/s. Despite the success of MC-IDS on HIT-SI, a need for increased statistical significance, along with preponderance of the HIT-SI3 machine to operate in a lower light ( lower IDS signal ) regime, motivates an upgrade to a more sensitive, higher resolution setup. C-IDS achieves this by replacing the old fiber optical bundle with a new coherent one, with a tens of times more individual fibers. Results of comparison tests find that C-IDS has lower throughput than MC-IDS, demonstrating a tradeoff between temporal vs spatial resolution. Data collection methodologies and significant sources of error will also be presented.

Fetal Alcohol Spectrum Disorders (FASD) are considered to be the leading preventable cause of mental retardation throughout the world. Fetal Alcohol Syndrome (FAS), a specific syndrome under the umbrella of FASD, represents the extreme consequences of maternal alcohol consumption, which results in facial dysmorphologies and neurological defects. FAS is observed throughout the world and is difficult to quantitatively measure because phenotypic effects amongst races and ethnicities are often not identical. In this project, our lab is interested in assessing the extent to which craniofacial development of the skull and mandible is affected in offspring due to chronic maternal alcohol consumption during pregnancy by using a voluntary intake mouse model. Female mice were bred with males at the average age of 15 weeks, and were divided into three treatment groups based on the amount of alcohol introduced to their drinking water during their pregnancy term: the 10% ethanol/water group was exposed to the solution for the entire term of the pregnancy, the 15% ethanol/water group was exposed to the solution for the first 8 days of pregnancy, and finally there were water-only control groups. When litters are at 75 days old, the craniofacial phenotype of the samples are acquired via high-resolution microCT. Shape differences amongst the individuals will be statistically assessed using sets of 3D landmark that will be collected from the skull and the mandible using 3D Slicer software. These landmarks will be then analyzed using geometric morphometric methods (GMM). We are currently in the image acquisition and processing phase. As a whole, we expect to see subtle but significant differences in craniofacial development between mice that are born to mothers consuming alcohol during pregnancy and their controls. From there, the leading mechanism and degrees of alteration of craniofacial development can be related to the effects of prenatal alcohol exposure in humans.

The DELLA protein subfamily consists of five proteins found in Arabidopsis thaliana that function together with Gibberellin plant hormones to help regulate important aspects of plant growth and development. How DELLA proteins act to regulate transcription by repressing gibberellin to control the developmental processes of plants is still unknown. Preliminary structural studies were conducted on the C-terminal GRAS domain of RGL1, one of five proteins of the DELLA subfamily, to further examine its potential role in transcription regulation. The protein was expressed in Escherichia coli cells after using molecular cloning techniques to amplify and insert the protein sequence. Various purification processes were used to extract and purify the protein in preparation for crystallization. Crystal tray trials were then set up using the purified protein in hopes of obtaining a crystal structure of RGL1 for further structural studies. Also, previous sequence analysis of the GRAS domain show sequence homology to methyltransferases which are responsible for methylating their substrates. Determining the crystal structure and comparing with methyltransferases will be able to help clarify the potential function of the GRAS domain of DELLA proteins.

 Silver nanoparticles (AgNPs) have become an increasingly common component of many industrial products, causing concern over the potential for toxicity in both humans and in the environment. It has been proposed that a significant portion of these toxic effects may be due to the release of silver ions (Ag+) from the nanoparticles and subsequent damage through the generation of oxidative stress. To evaluate some of these concerns surrounding Ag+, the dose responses of two different transfected mouse hepatocyte cell lines were measured at incremental concentrations of solvated Ag+, delivered as silver acetate. Relative to the control HepaV line, CR17 cells have higher levels of the enzyme glutamate cysteine ligase (GCL), which catalyzes the first step in production of the antioxidant glutathione (GSH). Comparison between the two lines was done in an effort to establish a relationship between Ag+ toxicity and GSH-dependent defense against oxidative stress. Cell viability was measured using an MTT assay and GSH levels were measured using the naphthalene-2,3-dicarboxaldehyde fluorescence assay for total GSH. In addition, Western immunoblot analysis was conducted for the two subunits of the GCL holoenzyme, GCLC and GCLM. In all cells, a significant dose-dependent decline in viability was observed at 2 µg Ag+/mL and greater. However, CR17 cells consistently maintained a higher percentage of cell viability than did HepaV cells. Regarding GSH and GCLC/GCLM expression levels, significant variation in the two cell types was observed (as expected), and their levels appeared to increase with Ag+ dose. However, there was no significant dose-response observed. These results suggest, but do not conclusively prove, that oxidative stress is a mechanism for the observed toxicity of silver in these mouse hepatocyte cell lines. Future studies will focus on AgNP-induced oxidative stress and toxicity in these cells. 

Hydrazide glycosylation is a useful tool for the formation of bioconjugates such as oligosacchardies, glycopeptides, and other products. Hydrazide glycosylation reactions, however, do not proceed well in biological pH. With a goal of developing an efficient bioorthogonal neoglycosylation reaction, we investigated the formation of a glucosylhydrazide using various aniline derivatives as catalysts under acidid (pH 4.5) and neutral conditions (pH 7.4). Preliminary results show increased rates under acidic conditions and promising improvements under neutral conditions.

The nitrogen-vacancy (NV) center is a defect in diamond that has promising applications in quantum information processing based on quantum entanglement. A critical requirement for quantum entanglement is that separate NV centers have identical optical properties. Here we present the design and implementation of an optical characterization system for NV centers called photoluminescence excitation (PLE) spectroscopy. While scanning a NV center with light from a tunable-frequency red laser, we detect and analyze the intensity of the resulting photoluminescence. The suitability of this system for characterizing NV centers, resonantly exciting the centers, and providing feedback to stabilize the NV optical properties will be discussed.

The planet is experiencing dramatic climate shifts resulting in large regions enduring significant weather events of unprecedented proportions. Drought is a weather phenomenon that can cause devastating economic and agricultural impacts that many of these regions are unequipped to deal with. According to the population reference bureau (PRB), approximately half of the world’s population resides within 200 kilometers of a coastline. Thus, resorting to large bodies of water for clean drinking water is an obvious solution to unexpected drought. Zwitterions are environmentally friendly molecules that maintain a positive and negative charge simultaneously. Research has shown that this class of molecules possess non-fouling capabilities in a variety of marine applications due to their hydrophilic properties. Therefore, using zwitterionic polymer coatings in an effort to eliminate the membrane fouling issue that plagues reverse osmosis (RO) desalination plants is promising. Currently, the water is pretreated with harmful chemicals to reduce membrane fouling. Membrane fouling has prevented RO desalination from emerging as a viable source of fresh water due to the high maintenance and operation costs associated with this method. In the Jiang lab, I have synthesized zwitterionic polymers, poly(sulfobetaine methacrylate) (pSBMA) and poly(carboxybetaine methacrylate) (pCBMA), that have been grafted to the surface of RO membranes of varying compositions. The coated membranes were then exposed to solutions containing fluorescently tagged proteins. The non-fouling performance of the coated membranes was then compared to uncoated membranes by measuring the fluorescence intensity of the bound surface proteins. Additionally, enzyme-linked immunosorbent assays (ELISA) were used to quantify the surface protein density. Utilizing zwitterionic coatings to eliminate the fouling issues experienced by desalination plants will reduce the monetary and environmental costs of RO desalination. This action will further prevent harmful pretreatment chemicals from entering the environment.

Global energy consumption has reached an all-time high. In turn, there has been increased interest in university energy consumption in recent years, in hopes to reduce university related energy expenditures. Recently, reports estimating the sustainability of the UW Athletic Department were published in hopes to reduce University energy use and promote more sustainable practices. To further evaluate the athletic department energy usage, a model can be utilized to estimate total energy consumption for past or present years. The purpose of my research is to build a robust and versatile model that has the ability to estimate the energy expenditures of collegiate athletic programs. This model can be utilized for energy accounting and monitoring for sustainability initiatives. The model aims to be interactive, utilizing calculation parameters and inputs that can be easily modified by the user. Data characteristics and availability were examined in order to determine model structure and necessary calculations for energy accounting. The model structure was designed with two purposes in mind, usability of the model for the wide range of potential users and flexibility of the model for variations in data or calculation parameters. At completion, the model will calculate the energy expenditure and carbon dioxide production for varying types of athletic department functions, allowing for a diverse range of data and energy calculation parameters. Additionally, the model will be utilized to estimate the Athletic Department’s energy expenditure for the 2013-2014 Academic Year.

Chronic itch is a debilitating condition that plagues hundreds of millions of people. It is a symptom of many illnesses including cancer, kidney failure, liver cirrhosis, MS, and shingles. Finding treatments for chronic itch would greatly improve many peoples’ lives. The first step in developing effective treatments is to determine the cellular and molecular mechanisms responsible for chronic itch. This project aims to determine whether the itch transduction pathways identified in mammals are present in zebrafish (Danio rerio). Zebrafish are an excellent model organism. They reproduce in huge numbers and their nervous systems develop very quickly. Additionally, there are many powerful techniques available to study itch in zebrafish that are not possible in other model systems. The Dhaka Lab is interested in determining what forms of itch, if any, exist in zebrafish so that we may leverage the properties of the zebrafish model system to characterize the properties of itch stimuli transduction, thus expanding our knowledge of itch pathways and potentially gaining insight into the evolutionary origin of itch in vertebrates. We have been exploring whether compounds that induce itch, such as agonists of histamine receptor 1 and toll like receptor 7, also evoke an itch response in zebrafish. Our preliminary behavioral data shows that these compounds may induce itch in zebrafish. Additionally when fish were exposed to these compounds we observed activity in neurons of the trigeminal ganglion, an exclusively somatosensory cluster of neurons, which indicates that these compounds elicit a somatosensory response in zebrafish. We plan to continue characterizing these pathways to increase our understanding of itch, which might aid in the development of therapies to treat chronic itch.

Existing literature has established vicarious affect and reputational concern as two possible (but usually confounded) motivations for strategic prosocial behavior in children. The present research examines the types of situations in which children are more likely to engage in prosocial behavior. To disentangle these, we created a modified dictator game with a 2 (actor can see recipient) x 2 (actor can be seen by recipient) design to create conditions with only one motivation or the other present (addressing the vicarious affect and reputational theories, respectively). Children ages 4-6 were given the opportunity to choose how many M&Ms to give to themselves and to give to another child, with whom they were ostensibly interacting via video chat. In one condition, the video chat proceeded as planned, while in the other three, the camera "malfunctioned" on either the actor's end, the recipient's end, or both, so that the actor could experience being seen by the recipient separately from seeing them. We found that children were significantly more generous in the condition where they could both see and be seen by the recipient than they were in any of the other three conditions, none of which differed significantly from one other in terms of generosity rates. This finding suggests that the combination of these two motivations in some way leads children to be more generous than does either motivation individually. Current follow-up work attempts to parse out different facets of the combination condition of seeing and being seen, by separating it into the different components of social interaction and mutual knowledge of identities, in order to determine which qualities of the interactive experience contribute to children’s increased generosity.

During the critical period, a time beginning soon after birth and lasting weeks or months depending on species, visual experience can strengthen or weaken synapses in the visual part of the brain to optimize it for processing the stimuli that it sees. After the critical period ends, visual experience cannot change synapses. If it were possible to make a brain area plastic again, like it was during its critical period, it would enable experience to optimize the function of that part of the brain again. This could significantly aid rehabilitation after injury to the brain or body impairs function. Work in other labs show that injecting an enzyme that dissolves extracellular structures called perineuronal nets (PNNs) that form around neurons can restore critical period-like plasticity. PNNs are composed of several chondroitin sulfate proteoglycans (CSPGs) bound to a core protein, hyaluronan. Disease, trauma, and congenital problems with the system in the brain that moves the eyes can severely impair vision. We may be able to treat these problems by injecting the enzyme into oculomotor regions where neurons are surrounded by PNNs. To identify what parts of the oculomotor system contain many neurons surrounded by PNNs and therefore might be good injection targets, we detect PNNs by immunohistochemically staining Rhesus monkey brain tissue mounted on glass slides. I used wisteria floribunda agglutinin antibody, which binds to CSPGs, to mark PNNS. To mark neurons I used NeuN primary antibody followed by donkey anti-mouse secondary. We divide the number of neurons in an area by the number surrounded by PNNS to calculate the percentage of neurons surrounded by PNNs in a particular brain structure. Identifying nuclei with high percentages of neurons surrounded by PNNs will tell us where to inject our PNN-dissolving enzyme for maximum clinical benefit.

In spite of vast research relating to the foot, intrinsic foot musculature and the interrelation to the medial longitudinal arch (MLA) is not fully understood. The understanding of how intrinsic foot musculature is affected by everyday activities is necessary to prevent and/or treat chronic foot pathologies that may lead to injury. Discernment as to what causes the arch of the foot to become abnormal, as well as how to normalize and strengthen the active structures of the foot, can benefit individuals with poor foot posture. Therefore, the purpose of this study is to examine the difference in MLA and intrinsic foot strength between adult sedentary and non-sedentary populations. In order to fulfill the purpose of this study, the methods will consist of: specific background questions relating to lifestyle, Chippaux-Smirak index, sedentary lifestyle index (<5000 steps per day), body mass index, and a force platform configuration. The Chippaux-Smirak index is a reliable, valid, and effective tool to evaluate the medial longitudinal arch height via the use of a foot print. Also, a force platform configuration will be used to non-invasively test the strength of the intrinsic foot musculature. By the use of quantitative analysis, a resulting statistical significance will help differentiate unobservable disparities between the MLA and intrinsic foot strength among adult sedentary and non-sedentary populations. In addition, the background questions and body mass index will be used to describe any correlations, as well as confounding variables. The resulting data from this study may indicate a potential correlation between physical activity levels and arch height. In light of a potential correlation, this study will hopefully inspire medical practitioners to further stress the importance of physical exercise, with respect to foot health and overall wellbeing.

Autism Spectrum Disorder (ASD) describes a set of neurodevelopmental conditions characterized by behavioral and cognitive phenotypes with multiple gene involvement. There is a strong association between bowel disorders and ASD, making the enteric nervous system (ENS) a potential target for drug treatment and therapy. The ENS innervates the gastrointestinal system and regulates its function. Current studies of the ENS are limited to the invasive procedure of removal of gut tissue, or animal models that do a poor job of recapitulating the human disease. The work presented here uses induced pluripotent stem cells (iPSCs) from ASD patients that have been differentiated into enteric neurons to model the ENS for study. Fully differentiated enteric neurons pass through an intermediate cell type called neural crest (NC). Here, we report the successful differentiation of iPSCs to neural crest progenitor cells (NCPCs). Treatment of iPSCs with two inductive factors, SB431542 (a TGF-B Inhibitor) and BIO (a GSK-3 inhibitor) generated a high yield of NCPCs. Optimization of the protocol was accomplished by treating iPSCs with varied concentrations of BIO and SB to determine which combination yielded the highest percentage of NCPCs. The expression of NCPC markers p75 and Sox10 by immunocytochemistry, in the absence of Pax6 was used to quantitate the extent of NCPC differentiation. Together, these findings support the conclusion that iPSCs can be efficiently differentiated to neural crest progenitors as an intermediate in the generation of an “ENS in a dish” to study the chronic constipation suffered by patients with ASD.

One of the most widely used engineered nanoparticles—silver nanoparticles (AgNPs)—are being used increasingly in consumer products. AgNPs exhibit antimicrobial properties by disrupting cellular proteins, enzymes, and DNA. However, the adverse health effects of AgNP exposure are not well understood. The goal of this project is to evaluate the effects of AgNP exposure on early brain development. This is accomplished using 3D in vitro mouse micromass culture to observe the effects of AgNP on neuronal cells. This experiment uses C57BL/6 and A/J mouse strains to establish high density 3D micromass cell cultures. Mouse midbrain cells undergo numerous processes of neural development including proliferation and differentiation in vitro. Developing mouse midbrain cells are collected from fetuses at gestation day 12 for A/J mice and gestation day 11 for C57BL/6 mice. Using two mouse strains allows us to investigate the potential for genetic differences in response to test chemicals. Cells are plated and cultured for 1, 7, 14, and 21 days. Cultures at each of these timepoints are exposed to AgNPs and evaluated for toxicity. AgNPs of different sizes and coatings are tested including 20nm citrate coated AgNP (AgCitrate 20), 110nm citrate coated AgNP (AgCitrate 110), and 110nm PVP coated AgNP (AgPVP110). AgNPs are administered in four different concentrations: 6.25, 12.5, 25, and 50 µg/mL. In addition, positive and negative controls as well as citrate and PVP coating only controls are tested. Cytotoxicity is measured in all dose groups and controls by the presence of lactate dehydrogenase with an LDH assay at 24 hours after treatment. Evaluation of cytotoxic endpoints of the different AgNPs has allowed us to investigate the effect of the various AgNPs on developing neuronal cells. Results include development of a dose-response model of AgNP neurodevelopmental potential and will help to inform the growing field of nanotoxicology of AgNP effects on sensitive endpoints of toxicity.

Volunteering in America has been declining in recent years. Due to minimal community support, some non-profit organizations are struggling to obtain necessary items. To address this problem, our capstone project will create a platform to effectively communicate requests from Seattle non-profit organizations to local altruists. Our application, CherryTime, will enable organizations to make specific requests for items or volunteers. We will be developing a mobile-friendly website where locals will be able to fulfill these requests. Individuals will have the ability to look for opportunities based on location, time, causes they support or skills they have. Their donations and time volunteering will be rewarded through our badge system. Our project will help create a two-way relationship between organizations and individuals. The goals of this project are to increase the number of donors and volunteers in Seattle, make it easier for non-profit organizations to communicate their needs and to raise awareness for charity groups and their needs.

Annually, over 300,000 babies worldwide are born with neural tube defects (NTDs) that can lead to death or significant neurological pathology. The main causes of NTDs are genetic mutations of the planar cell polarity (PCP) pathway, the process by which cells are correctly oriented and localized in a 2D plane during development. We can visualize disruptions of the PCP by observing phenotypic processes including the blocked migration of facial branchiomotor neurons (FBMNs), shortened body length, and disruption of floor plate polarization (FPP). We conducted a reverse genetic screen on Danio rerio (zebrafish) utilizing the CRISPR/Cas9 system, which is an effective genetic engineering tool. We hypothesized that there are core genes such as VANGL2 and SCRIB involved in some, but not all of the PCP phenotypic processes. LPP knockdown disrupts both convergent extension (CE) and FPP. SEC24B knockdown affects CE. Both SEC24B and LPP do not disturb FBMN migration. Therefore, different phenotypic processes recruit specific genes. This research may help elucidate the intricacies of the PCP pathway and identify genes central to NTDs, allowing for development of gene-specific therapeutics.

Of all the crimes commonly investigated by law enforcement, arson is one of the most difficult to prosecute. One piece of key evidence is the identification of accelerants, substances used to either speed up the fire or make it burn hotter. Most of the original material is consumed in the fire, leaving only traces to be analyzed. It is the purpose of this study to look at the detectability of these traces under different conditions common to arson cases. This research examines the capability of a portable miniature gas chromatogram (mini-GC) to detect and accurately identify acetone, paint thinner, and denatured alcohol. Experimentally, this is done by burning several types of plastics, carpeting, and wood using these accelerants. Due to the nature of the mini-chromatogram only the lower molecular weight compounds (C₂ – C₈) are tested. The burned materials are sampled using headspace solid-phase micro-extraction (HS-SPME), a passive sampling method common to the forensics field. The samples are analyzed by the mini-GC with a chemicapacitor sensor and later with a more sophisticated gas chromatogram mass spectrometer (GC-MS) to verify the findings. Unburned and burned samples without accelerant are also sampled and analyzed as a control. The resulting chromatographs are reviewed and compared to chromatographs of the pure accelerants to determine whether or not acceptable peaks of the accelerant traces are visible. This research could be useful to investigators in determining the viability of the mini-GC as a forensic tool. Further work can be done to optimize chromatogram settings for different accelerants and examine the detectability of larger carbon compounds with the GC-MS. This is done with the intention of sharing the findings with the Washington State Patrol Crime Lab.

The death of a parent is a traumatic event in children’s lives. It may lead to posttraumatic stress (PTS) symptoms and maladaptive grief in children, as well as depression, anxiety, anger, hopelessness, and suicidality. Sub-Saharan Africa has a particularly large orphan population, as it is home to 90% of the 16.6 million children orphaned by HIV/AIDS around the world. For the present study, researchers from the University of Washington and other U.S. universities partnered with the Tanzania Women Research Foundation (TAWREF) in Tanzania to address this problem. The intervention was group-based, culturally adapted Trauma-Focused Cognitive-Behavioral Therapy (TF-CBT) delivered by trained lay counselors across 12 weekly sessions. Participants were orphaned children (7-13 years old, N = 64) suffering from symptoms of maladaptive grief and/or PTS, and their guardians. Guardians received the same intervention as children, as well as additional components to help better support the child. Outcomes of children’s grief, PTS, behavioral and emotional well-being, and depression were measured at four time points: baseline, end of treatment, 3 months follow-up, and 12 months follow-up. We examined how well child-guardian relationship quality predicted children’s treatment outcomes, as well as how child-guardian relationship changed over the course of the intervention. If results prove the importance of strong child-guardian relationship for orphans’ outcomes and that the intervention serves to improve that relationship, this would provide strong support for the use of TF-CBT as a therapy that requires strong guardian involvement and commitment.

Adverse drug reactions (ADRs) are defined as “an appreciably harmful or unpleasant reaction, resulting from an intervention related to the use of a medication”. They can cause harm to the patient and are also responsible for increases in medical cost. Many ADRs are a consequence of drug-drug interactions (DDIs) that promote alterations in drug metabolism. In fact, evidence suggests that the majority of DDIs are mediated through induction or inhibition of Cytochrome P450 enzymes isoforms (P450). Therefore, understanding P450 mediated DDI and their impact on drug metabolism is critical in preventing ADRs in humans. Proper tools are needed to identify potential DDIs and investigate particular CYP isoforms’ metabolism. Chloromethiazole (CMT) is a FDA acceptable inhibitor of CYP2E1 for in vitro studies; however, little is known about its utility as probe inhibitor. We are interested in determining the mechanism and selectivity of CYP2E1 inhibition by CMT; we also seek to demonstrate its utility as an inhibitor of acetaminophen bioactivation by CYP2E1, which is relevant to treatment of overdose.
To understand CMT’s inhibitory mechanism, we will synthesize deuterated analogs of CMT and to elucidate its mechanism of irreversible enzyme inactivation via thiazole bioactivation. Replacement of selected hydrogen atoms with deuterium should help to identify metabolic sites that are responsible for inactivation due to changes in the inactivation rate arising from kinetic isotope effects at these metabolic sites. Further, selectivity of inhibition will be assessed for irreversible and reversible inhibition of major P450s by experiments with accepted probe substrate and LC/MS/MS analysis. If the data support these hypotheses, we can test the drug with cryopreserved human hepatocytes as a model system to assess acetaminophen toxicity. The completion of this project will help characterize CMT as a selective CYP2E1 inhibitor and as a potentially beneficial drug to treat CYP2E1-mediated acetaminophen toxicity.

Despite women's increased labor force participation in the United States since the 1950's, women continue to perform a significantly greater amount of household work than men. The gender gap is particularly pronounced among married individuals. Women who work full-time and are also responsible for the majority of household work are said to bear the burden of the "second shift". An existing body of literature examines the role of bargaining power in household decision-making processes and the division of household work. My research is contextualized within this literature and asks: What demographic factors determine the amount of time women and men spend performing household work? Do the determinants of time spent in household work influence women differently than men? I use data from the 2013 American Time Use Survey to examine how education, race, and family structure impact the number of hours women spend performing household work in comparison to those spent by men. After controlling for other relevant characteristics, education level, race, and family structure are significantly related to time spent in household work. The magnitude of these relationships varies by gender. 

Tuberculosis (TB) kills approximately 2 million individuals worldwide annually. Toll-like Receptors (TLR) and the Interleukin -1 Receptor (IL-1R) are critical mediators of effective macrophage immune response to tuberculosis. The IL-1 and IL-1R signaling system includes three cytokines that bind IL-1R: IL-1α, IL-1β, and interleukin receptor antagonist (IL-1Ra.) IL-1R associates with Interleukin Receptor Associated Kinase 1 (IRAK-1) though its Toll/Interleukin Receptor (TIR) domain, which ultimately leads to the induction of inflammatory genes. TLRs also contain a TIR domain and associates with IRAK-1, but their ligands are microbial products. Common variations in one TLR regulatory protein, TOLLIP (Toll Interacting Protein), are associated with susceptibility to TB. TOLLIP inhibits IRAK-1 and may regulate both TLR and IL-1R signaling pathways. However, it is not known whether TOLLIP regulates Mycobacterium tuberculosis (MTb)-induced cytokine production or affects MTb replication within cells. We hypothesized that TOLLIP diminishes IL-1R signaling in response to IL-1β, and alters MTb replication in monocytes. To test this hypothesis, we created TOLLIP-deficient cells in the THP-1 human monocyte cell line. We stimulated cells with either IL-1β, LPS (TLR4 ligand), or MTb whole cell lysate (ligand for multiple receptors including TLR2) and used ELISA to measure secretion of IL-6, IL-1Ra, and TNF. TOLLIP-knockout cells secreted less IL-1Ra compared to controls after IL-1β (p = 0.0039, Students’ t-test), LPS (p=0.029), and MTb whole cell lysate (p=0.019) stimulation. In separate experiments cells were infected with live MTb. MTb replication was reduced in TOLLIP-deficient compared to control cells (p=0.001). This data suggests that TOLLIP regulates IL-1Ra and IL-1β secretion after signaling through both the IL-1R and TLR pathways. Future experiments will focus on determining the mechanisms by which TOLLIP influences IL-1 receptor signaling. Knowledge of how TOLLIP shapes the immune response to infection could be harnessed for the development of host-directed treatments and vaccines for diseases like leprosy and TB.

Complications associated with premature birth often disrupt normal brain development and/or cause direct damage to the developing brain, resulting in poor neurodevelopmental outcomes. Physiologically relevant animal models of perinatal brain injury can advance our understanding of these influences and thereby provide opportunities to develop therapies and improve long-term outcomes. While there are advantages to currently available small animal models, there are also significant limitations that impede the effective translation of research findings to humans. Large animal models (piglet, sheep, nonhuman primate) have complex brain development more similar to humans, but these animals are expensive, have longer periods of gestation, and often require complicated husbandry. Ferrets (Mustela putorius furo) are born lissencephalic, or with a smooth brain, and undergo postnatal cortical folding to form complex gyrencephalic brains like those seen in larger animal models and humans. This literature review examines the use of ferrets as models of human disease, and describes other attributes of ferret brain growth and development that make it an appealing animal in which to model perinatal brain injury. We postulate that because of their innate characteristics, ferrets have great potential in neonatal neurodevelopmental studies.

The word "indigenous" continues to evoke discussion and scholarship throughout the ages as a highly culturally and politically loaded term that has gained global importance. The Islands of Mauritius, Réunion and the Seychelles in the Indian Ocean are fascinating examples of the changing uses of the notion of “indigeneity” because although there are no known “indigenous people” that resided on the land, there is still a distinct hierarchy of privilege that continues to reinforce colonial ideals of indigeneity onto those of Creole or African backgrounds. How do Creole peoples view themselves? Do they see Creole identities as “more indigenous” than Indian or White settlers? How does their relationship to a complex Métissage (mixed heritage) relate to global discourse of indigeneity? In this paper, I plan to use the above island nations to explore notions of indigeneity in a global context and to understand how many residents define themselves within this framework. I argue that the indigenous-settler binary is deeply rooted in colonial discourses and that it has not simply disappeared but evolved through the legacy of Creole people (Métissage) and the complex relationship that they have with France, their former colonizer.

The objective of this project was to detect if the pollutant levels in the city were significantly higher than pollutant levels in a rural setting. A place that’s more densely populated would be expected to have more traffic concentration, and therefore higher pollutant concentrations. The hypothesis for this project was that the city would yield higher pollution readings during high traffic periods compared to the rural pollution readings during high traffic periods. Carbon dioxide and nitrogen dioxide exposure during bus commutes was measured using an Arduino platform. The sensors used were the TGS2442 and MiCS-2710 for carbon monoxide and nitrogen dioxide, respectively. One of the starting points tested bus commute pollution exposure from Monroe and compared it to the bus commute starting from Seattle. The two separate starting points and paths followed to the same end point, Bellevue College. The tests were performed at different times of the day where traffic was predicted be at its peak volume (based on WSDOT reports), and during periods of low traffic volume to explore any other trends in pollution levels. No significant disparity resulted between carbon monoxide exposure between comparison of the two starting locations to the final location. However, nitrogen dioxide levels did spike for the rural readings, most likely due to passing by manure farms. Between high traffic volume time periods and low traffic volume time periods, however, the former showed a higher qualitative trend in pollution exposure where the latter showed lower levels of exposure to air pollution. In other words, this could imply that the time of day has a greater impact on how much pollution a commuter is exposed to, not necessarily how densely populated an area is. This could also imply that a health-conscious commuter should consider the time of day they start their commute over location.

Urban community garden programs across the United States have had numerous positive impacts on health and socio-economic factors. Seattle is one of the leading cities in the United States for its number of urban community gardens. This research examines the relationship between Seattle's "P-Patch" gardens and crime, with a particular focus on the presence as well as the perception of neighborhood crime. We utilize quantitative, qualitative, and spatio-temporal GIS methods. A geographic information system (GIS) is used to map and analyze 84 P-Patches throughout Seattle, crimes in 1996 and 2006, and socio-demographic data. Bivariate and multivariate analyses are used to examine any positive or negative correlation between crime and P-Patch presence. This preliminary research will be further developed with an advanced GIS analysis (e.g. crime hot-spot analysis) and iterative qualitative analyses including interviews with P-Patch gardeners to observe their perceptions of the relationship between P-Patch presence and crime. We hope to contribute to existing studies on community, urban community gardens, urban crimes, as well as local politics around the P-Patch community garden program. 

This project evaluates the efficacy of coatings, containing gallium-metal complexes versus an antibiotic (gentamycin), deposited on orthopedic metal implants to inhibit the growth of bacterial biofilms and therefore reduce the occurrence of post-surgical infection. Biofilms account for a majority of hospital acquired infections (4^th leading cause of death in the US), decreasing the rate of occurrence would also increase the recovery rate of patients, as well as reduce health care costs. Hypothetically, by increasing the concentration of gallium complexes and gentamycin in the metal coatings, we will observe a significant decrease in biofilm formation. Here, we cultivated the bacterial species, Pseudomonas aeruginosa and Staphylococcus epidermidis in batch suspended cultures, in which the coated discs were submerged. Discs were exposed to bacterial cultures for 48 hours, with liquid samples taken periodically to determine live/dead cell concentrations. Adherent cells on each disc were determined after 48 hrs by sonicating the disks to remove any adherent bacterial growth; live/dead cell counts as well as agar plate (CFUs) counts were carried out. Preliminary results thus far indicate that higher levels of gallium complex and gentamycin were able to significantly reduce bacterial colonization. Experiments are still on-going.

Malaria is a parasite-borne illness that causes about 600 million deaths annually. Reports of drug resistance to frontline antimalarials in Southeast Asia highlight the pressing need to develop new drugs that will be potent against existing drug-sensitive and drug-resistant Plasmodium species of malaria parasites. A clearer understanding of how easily parasite resistance occurs against experimental antimalarials may lead to better treatment designs, as well predictions of how well the drug will perform when administered clinically. Pyronaridine is an antimalarial drug used mainly in China for the past 30 years, but has recently been approved in Cambodia and is a candidate for even more widespread use. The mechanism by which pyronaridine inhibits the parasite is not well understood. One approach to identifying the mechanism of resistance to pyronaridine is to select for resistance against whole cells. Using the species infecting humans, Plasmodium falciparum, we attempted continuous and step-wise increasing drug pressure schemes to generate resistance in vitro. Preliminary results indicate that low-level resistance to pyronaridine can be obtained using the step-wise method of selection. Additional selections may yield higher-level mutants. Mutants obtained from these studies will then have their genomes sequenced and compared to sensitive parasites. From this we may identify the mechanism of resistance to pyronaridine, which could help in understanding and predicting the efficacy of pyronaridine use in the clinic.

Recent spinal cord injury studies have focused on recovery of limb function after incomplete contusion injuries, where some axonal fibers are spared following an injury. In this study we are exploring methods to strengthen the remaining axonal connections following injury via mechanisms of Hebbian plasticity, in which repeated stimulation of postsynaptic neurons gated by presynaptic activity increases the synaptic strength between pre and postsynaptic neurons. After receiving a cervical level lateralized contusion injury, rats are implanted with four electrodes including spinal surface electrodes both rostral and caudal to injury, an array of intraspinal electrodes caudal to injury, and electromyogram (EMG) electrodes in the muscles of the paralyzed forelimb. The rats are then tested before and after a treatment period comprising of activity-dependent intraspinal microstimulation (ISMS). During this treatment phase, the spinal surface electrodes are used to record electrical activity on the surface of the spinal cord and subsequently trigger ISMS. During the test phase, stimuli are delivered to the spinal cord surface and the successive evoked potentials are recorded using the intraspinal electrodes and the EMG electrodes. We are hopeful that the activity-dependent ISMS delivered during the treatment phase can induce timing dependent (Hebbian plasticity) among the remaining axonal connections that bypass the injury.

The structure of a protein has long been associated with its function. Due to both divergent and convergent evolution, evidence exists to indicate not all proteins with similar structures share similar functions. However, there is interest in whether these similar structures share similar dynamic behaviors. Proteins that share similar 3D structures, or folds, can be grouped into folding families, and previous work suggests that there may be common mechanisms by which these proteins fold within these folding families. Here, we investigate the dynamics and unfolding pathways of 17 members selected from the sixth most populated fold family, the SH3-like barrel domain. These proteins have a wide range of functions including cell signaling, protein-protein interactions, muscle protein function, and RNA regulation. The SH3-like barrel proteins have been the target of many previous studies, and consist of small, β-strand proteins. Using molecular dynamics simulations at high temperatures, the unfolding pathways, including the characterization of the transition states, are studied for each of these proteins. With a broad analysis of native and non-native contacts as well as secondary structure content, common folding patterns have been seen among this folding family. Comparative analysis of our molecular dynamics simulations to the findings of others confirms a hierarchical pattern of contact loss. Reported here are the conclusions of our analysis of the unfolding pathways of 17 members of the SH3-like barrel domain, including the commonalities and differences of these pathways to those found by others.

The abiotic (before life) formation of biopolymers (specifically DNA, RNA, and proteins) has long been a mystery and is important for understanding the origin of life on earth. These bio-molecules are composed of building blocks that would have been dispersed in early oceans. Our previous work has shown that RNA bases and ribose bind to and stabilize fatty acid vesicles. Our results implied the building blocks of a biological polymer could have spontaneously associated with components of the first membranes to form stable structures. We have now shown that protein building blocks, too, stabilize fatty acid vesicles against salt induced flocculation (a type of aggregation in which the vesicles come out of suspension). Using spectrophotometry, we measured the presence of flocs (and other structures) in fatty acid solutions, with and without amino acids over a range of temperatures. Using fluorescence microscopy, we identified the structures that caused changes in absorbance in our spectrophotometric assays. We found that the two most hydrophobic amino acids, inhibit salt induced flocculation. Moreover, although alanine and glycine, which are less hydrophobic, had little effect on flocculation, dipeptides composed of these amino acids preserved vesicles in the presence of salt at 60 degrees C. These vesicles appear to be primarily multilamellar (composed of multiple bilayers) structures, which may promote reactions between components of biopolymers more effectively than unilamellar vesicles (composed of one bilayer). Thus prebiotic membranes could have facilitated the formation of peptides by bringing amino acids together, and peptides could have increased the formation of stable membranes. Such an auto-amplifying system, combined with selection for more effective peptides, could have led to the first cells.

Created as a platform to begin discussion regarding water crises and how to address them, Inspired, Low-Cost Methods of Purifying Water is a technological examination of unique systems that can be used by small, resource-limited communities to alleviate the impact of physical and economic water scarcity. Our article is a shortlist of cleverly-crafted, atypical, low-cost, efficient methods of redeeming water. We crafted this inventory by considering over 45 separate methods and selecting applicable techniques based on their relative levels of feasibility, efficiency, sterility, approximate cost, and environmental impact. By generating unusual alternatives to water purification, we hope to provide an opportunity for the global community to discuss and implement new projects concerning the alleviation of public health issues regarding access to water, improving local productivity in agricultural systems, and resource sustainability.

Giardia lamblia is a protozoan parasite most known for being the causative agent of giardiasis, a disease characterized by Giardia’s infectious colonization of the small intestine. With many of its cellular pathways containing fewer components than those of other eukaryotes, Giardia is also a minimalist model organism. The binucleate protozoan undergoes cytokinesis, for instance, without the actin-binding motor protein myosin, which drives membrane constriction in most other eukaryotes. In some eukaryotes, however, cytokinesis can still proceed after depleting or poisoning myosin motors. This suggests there is a non-myosin-based mechanism underlying cytokinesis. Therefore, the study of Giardia cytokinesis offers the possibility of elucidating the core components required for mechanical separation of cells during cytokinesis. In the case that Giardia uses novel cellular components for cytokinesis, the potential to identify novel therapeutic targets also exists. In order to shed light on Giardia’s cytokinetic mechanism, we are investigating how actin contributes to myosin-independent cytokinesis. Localization of actin by immunofluorescence microscopy has revealed that actin localizes to the leading edge of the cleavage furrow. Depletion of actin with translation-blocking morpholinos results in delayed cytokinesis and failed division events. Our working hypothesis is that actin plays a role in the membrane remodeling required for cytokinesis. To test this hypothesis, we have inhibited membrane remodeling with brefeldin A (BFA, an ER-to-Golgi trafficking inhibitor). This has resulted in similar defects to our actin-depletion results, thereby supporting a case for actin’s role in membrane remodeling during cytokinesis. We are continuing to pursue the role of conserved membrane trafficking components, which have been shown to be required for cytokinesis in both plants and animals. Our goal is to determine if Giardia may be a model for the underlying mechanism of cytokinesis that takes over in myosin-depleted cells.

In silicon photonics, light is propagated through narrow silicon waveguides at the nanoscale. Confined to a microring waveguide structure, this light produces an electromagnetic field that extends beyond the ring. The shift in peak resonance frequency of silicon microring resonators is monitored in real-time to determine the binding of biological analytes onto the sensor surface. Combined with a microfluidic platform for directing a flow over the microring resonators, silicon photonics offer a highly sensitive method of determining the concentrations and binding activities of biological analytes of interest in serologic assays. Protein fouling, the non-specific accumulation of protein molecules on a surface, presents a significant challenge in biosensing in biological media. Our lab has aimed to significantly reduce non-specific adsorption by coating a biosensor surface with zwitterionic polymer chains, resulting in a hydrated surface which resists protein fouling. We utilize poly(carboxybetaine) (pCB) coatings on silicon photonic microring resonators to minimize fouling during microfluidic flow of complex biological media. The conventional process of “growing” this polymer layer occurs by atom-transfer radical polymerization (ATRP) under an oxygen-free environment. The Jiang Lab of the UW Department of Chemical Engineering has developed a method for ATRP in the presence of air, known as activators regenerated by electron transfer (ARGET) ATRP. In collaboration with the Jiang Lab, we implement ARGET ATRP growing pCB coatings on our silicon microring biosensors. The reaction conditions were optimized for non-fouling coatings on the microring resonator platform, and non-fouling properties were observed following flow of whole plasma via a microfluidic platform. The pCB coating grown by ARGET ATRP was functionalized by conjugation of biological capture elements, and we show specific capture of biological analyte. This is a novel implementation of ARGET ATRP on silicon substrates, resulting in polymer coatings that improve non-fouling properties of biosensing silicon photonic microring resonators.

Giardia is one of the most evolutionarily divergent protozoans identified to date, and is thought to belong to an ancient lineage known as the Excavates. One of its many defining characteristics is the absence of myosin, the motor that drives membrane constriction during cell division or cytokinesis in nearly all eukaryotes. To study how Giardia performs cytokinesis without myosin, we are investigating the role of the membrane remodeling protein Rab11. Rab11, a G-protein that regulates membrane trafficking, has been identified as an essential component of the division machinery in other eukaryotes. We have begun characterizing the role of Rab11 in Giardia cytokinesis by performing immunofluorescence microscopy in order to determine its localization throughout the cell cycle. Furthermore, we are testing Tet-inducible dominant negative and constitutively active Rab11 mutants for a role in cytokinesis. With these results, we can determine the importance of Rab11 in Giardia cytokinesis. If it has an evolutionarily conserved role, Giardia may prove useful as a model for myosin-independent cytokinesis.

I am doing research on how the microscopic parasite Cryptosporidium impacts a small rural population in the city of Viçosa, Minas Gerais, Brazil. We are currently doing a cross-sectional pilot study to assess the feasibility of doing a larger study in the area. This work will bring together researchers from the UW (Seattle), Washington State University (Pullman), and Universidade Federal de Viçosa. Cryptosporidium mainly causes diarrhea, and sometimes leads to death. There are multiple Cryptosporidium subgroups, each relating mostly to a single animal species, but many times can be present in other species. For example, the subgroup Cryptosporidium parvum comes from cattle, but it is also found in humans. We are sampling water, humans and cattle and identifying subgroups of Cryptosporidium present in each of them. My literature review showed that there is little scientific information regarding which subgroup mainly contaminates Cryptosporidium-infected areas in Brazil. The study population in Viçosa is small and the area is agricultural, so I chose to sample water, humans, and livestock to show whether there is variation of the parasite according to the different sources. To be able to do human sampling, I filled out an extensive IRB application and we are now waiting for the approval in order to start the sampling. Sampling and analysis for this study is projected to start in September 2015.

The ram accelerator is a chemical mass driver that accelerates projectiles up to hypersonic velocities. The ram accelerator accomplishes this via ramjet-like propulsion cycles through a tube filled with gaseous propellant. The majority of ram accelerators built around the world utilize a smooth inner bore in the launch tube. However, this design has limits on the heat release of the propellant, which limits the peak thrust that can be generated at any given fill pressure. The baffled tube ram accelerator surpasses this limitation by replacing the smooth bore with a tube contoured with internal baffles. The introduction of baffles into the launch tube enables the use of propellants with higher heat release, enabling greater thrust and more efficient performance. Additionally, the projectile acts as a “one-way” valve as it passes through the baffles, allowing propellant intake necessary for the ram effect, but prevents blow-by which can rapidly decelerate the projectile. A proof-of-concept prototype was built and successfully tested at the University of Washington from 2001-2005, yet was unable to determine limits of operation. The goal of this project is to design, install, and test a 4-meter long test section in order to empirically deduce the limits of operation of the baffled tube ram accelerator. The project is currently in the production phase. I have been using SolidWorks 3D modeling and ANSYS finite element analysis tools to design an assembly that balances durability, functionality, and cost before it is manufactured. After the tube is built and installed, testing will begin. Should the baffled tube design yield higher thrust and more efficient propulsion than its smooth bore predecessor, then the baffled tube ram accelerator may be used in a variety of direct space launch, deep core drilling, and strategic defense applications.

Hearing loss can be caused by a variety of events. Aminoglycoside antibiotics, some cancer drugs, exposure to loud noise, and aging can all damage and kill hair cells irreversibly in the mammalian inner ear. Hair cells, the sensory receptors of the mammalian inner ear, consist of the cell body, short actin-based stereocilia, and longer microtubule-based structures called kinocilia. In the mammalian inner ear hair cells are fundamental for hearing, as well as balance. Therefore, hair cell death can lead to deafness and balance disorders. Hair cells, and support cells clustered into structures called neuromasts, make up the lateral line, a sensory organ, in fish. The lateral line allows fish to sense movement in the surrounding water, and is important in schooling behavior. Unlike mammals, however, fish (e.g. zebrafish) are able to regenerate lost hair cells. The 1060 mutation was identified through a genetic screen for resistance to neomycin induced hair cell death in zebrafish. Through genetic mapping, the mutation that causes resistance has been narrowed to one end of chromosome 24. Going forward using a derived Cleaved Amplified Polymorphic Sequences assay, which allows for the identification of single nucleotide polymorphisms within genes located in the genomic region, will allow for the specific determination of the mutated gene. Future work will concentrate on characterizing how the mutation confers resistance to antibiotics. We hope to gain new insight into cellular mechanisms behind hair cell death to protect people from unintended hearing loss.

Tauopathies are a category of diseases characterized by a direct link between neurodegeneration and the dysfunction of a microtubule binding protein called tau. In its native form, tau stabilizes microtubules, the “highways of the cell”, and helps regulate the transport of important cargo within nerve cells. In tauopathies such as Alzheimer’s disease (AD) and chronic traumatic encephalopathy (CTE), tau will dissociate from the microtubule and form characteristic dense aggregates of protein called neurofibrillary tangles (NFTs). Tau aggregate formation correlates with, and appears to contribute to, the progression of these diseases, leading to intense interest in therapeutic strategies focusing on tau. However, little is known about the mechanism of tau aggregate formation and its inhibition – a fundamentally important piece of information when considering tau as a therapeutic target. In order to shed some light on this question, we are studying how selected small molecules can interact with tau in vitro to either inhibit or facilitate aggregation. These compounds include a set of natural products thought to interact with tau, as well as widely used pharmaceuticals that correlate with either increased or decreased risk of dementia in population studies. In order to study the effects of these compounds on the aggregation of tau, I performed a series of fluorescence assays in which aggregate formation was monitored, over time, in the presence of selected concentrations of each compound. I then analyzed the data from these experiments to extract information about how the compounds altered the rates and mechanism of tau aggregation. This work aims to generate a detailed and mechanistic understanding of pathological tau aggregation. It also provides a foundation for identifying and analyzing the activity of other small molecules, potentially bringing us closer to untangling the mystery of Alzheimer’s and other equally devastating neurodegenerative diseases.

The Type Six Secretion System (T6SS) is a bacterial toxin-delivery system targeting bacterial cells which neighbor the donor. By understanding the T6SS, we can potentially treat biofilm related problems more efficiently by implementing it into medicine. The current model of this system suggests that it is structurally similar to a bacteriophage and injects effectors into a neighboring bacterial cell to promote the neighboring cell’s death. T6SS is a complex molecular machine made out of at least 15 proteins found in an abundance of bacterial species. We test the current model of T6SS in vivo by using fluorescence microscopy to quantify the dynamics of two essential proteins of the system, Fha and ClpV, in P. aeruginosa (PA). Fha is a membrane protein hypothesized to be one of the first proteins to interact with the structure’s site, while ClpV is hypothesized to disassemble the structure after the system is fired. We also measure how proximity of PA to B. thailandensis, an antagonistic bacterial species which also has an active T6SS, affects the dynamics of these protein interactions. We expect Fha and ClpV to be more active as the proximity of PA to B. thailandensis decreases.

Helping behaviors in adults can be influenced by many situational factors, such as the intentions of the individual requiring help, or that individual’s previous behaviors. However, there is continued debate over whether children engage in similar selective helping behaviors. This study examined whether 3-year-olds’ helping behaviors are influenced by the valence of an individual’s goals. Children were given the opportunity to provide a requested functional or dysfunctional object to an experimenter with either a neutral or a mean goal. Functional objects were appropriate for completing a particular task, while dysfunctional objects were not. For instance, one functional object was a cup for holding juice, while the paired dysfunctional object was a cup with a hole in the bottom. Neutral goals were tasks such as hiding a bear that was a surprise for a friend, while the equivalent mean goal was to hide the bear because it was stolen from someone else. We predicted that when the experimenter had a neutral goal, children would provide the requested object significantly more frequently when a functional object was requested than when a dysfunctional object was requested, but that children would provide the requested object more often in mean-dysfunctional trials than in neutral-dysfunctional trials. Our results showed that children were significantly more likely to provide the requested object when the experimenter requested a functional object than when the experimenter requested a dysfunctional object. However, for dysfunctional trials children were not significantly more likely to provide the requested object in mean trials than in neutral trials. This suggests that children behave paternalistically by providing a functional object even when a dysfunctional object is requested, but that children do not differentiate between neutral and mean goals when determining which object to provide.

End stage renal disease (ERSD) is a serious disease currently affecting over 871,000 people in the US. ERSD is the result of chronic kidney disease which can be caused by diseases of podocyte loss such as focal segmental glomerulosclerosis (FSGS). Treatments include immunosuppressants and glucocorticoids that are unspecific, long lasting, and have unwanted side effects such as osteoporosis and diabetes. This problem occurs when drugs that are meant to restore podocyte functionality affect other cells throughout the body. These diseases damage podocytes, thus making podocytes the ideal receptors for targeted therapy. Drugs conjugated with targeting peptides are an attractive solution in that the selected targeting peptide sequence binds to certain proteins and as a result, can achieve high specificity as well as reduce unwanted side effects. The method used for finding binding peptides is through a process called phage display. In phage display, a podocyte protein is immobilized to a well and exposed to a peptide-expressing library of M13 bacteriophages. Phage display was performed on three proteins, NEPH1, nephrin, and SMPDL-3b. NEPH1 and nephrin were selected because they play an important role in podocyte adhesion. SMPDL-3b was chosen due to its relationship with rituximab, a chimeric monoclonal antibody that has shown to regain podocyte structure by binding to SMPDL-3b. Rituximab was used to elute out possible peptides of interest. After 4 rounds of screening, the phage libraries were analyzed by next generation Illumina sequencing to identify potential peptide binders. By conducting phage display to find potential binders and analyzing prevalent motifs by Illumina sequencing, a peptide ligand that is highly selective could be discovered.

The northern kelp crab (Pugettia producta) is one of the most common species of crabs found on the west coast of the United States. Despite its large size, ecological importance, and abundance, nothing is known about its larval development. I am in the process of completing illustrations and a description of the first zoeal stage of this species, and will be comparing it to larvae of the graceful kelp crab (Pugettia gracilis), the only other member of the genus on the west coast for which a larval description exists. Describing distinctive features and characteristics of the northern kelp crab larvae will greatly aid in future diet analyses of fish and other predators, and in general studies of plankton along the west coast.

During body axis formation in zebrafish, a population of undifferentiated progenitor cells at the posterior end of the zebrafish embryo contributes cells to the trunk and tail. Previous research in our lab shows that the posterior progenitors spend little time in the G1 phase of the cell cycle, and instead are predominantly in the S and G2 phase, and enter a period of quiescence before differentiation. A short G1 is a hallmark of embryonic stem cells in vitro, and we set out to investigate whether a prolonged G1 phase inhibits differentiation of progenitor cells in vivo. In order to induce a G1 cell-cycle arrest in developing zebrafish embryos, we developed a transgenic zebrafish line which carries a dominant negative version of cyclin D1 under the control of a heat shock promoter. Upon activation, the cyclin D1-CDK complex, which is critical for G1/S entry, can be inhibited as a result of the expression of the transgene. We found that blood circulation is lost in a portion of the transgenic zebrafish embryos, while the number of blood cells was unchanged. These results suggest that extending the G1 phase in early embryos inhibits vasculogenesis. Embryos which express the transgene also have significantly thinner trunks than wild-type embryos. Muscle, which contributes to the girth of the trunk and tail, and vascular cells grow from a common population of progenitors in early embryos, and we are currently investigating what connection narrow trunks might have to defective circulation in embryos with a prolonged G1 phase.

Through my experience as a non-immigrant visa holder who grew up in the U.S. since elementary school, I realized that visa holders could also experience vulnerabilities and struggles as other groups of immigrants. However, I also learned that they are often invisible in immigration discourse. This honors thesis employs interviews of non-immigrant visa students (F1, E2, H4) who grew up in the U.S. since an early age, to make visible of the vulnerabilities in their life experiences as well as compare and contrast their experiences with other immigrants in the U.S. Interviewees’ responses show that non-immigrant visa students experience vulnerabilities and struggles, such as identity crisis, limited mobility and agency through restrictions on work and travel, fear of losing legal status, and financial as well as emotional hardships. My analyses also demonstrate how students navigate the realm of temporary status to continuously remain in the U.S. Overall, this study shows that even non-immigrant visa holders who are often invisible to society and sometimes seen as immune to struggles due to their race, ethnicity, and legal status, can experience similar and different vulnerabilities as other immigrant groups in the U.S., such as undocumented immigrants or permanent residents. I hope that this study will contribute to increasing the visibility of and advocacy for the non-immigrant visa holders in the U.S.

Mild traumatic brain injury (mTBI) caused by repetitive explosive exposure is a very common injury that Iraq veterans experience. Although it may not cause paralysis/death, it has devastating immediate effects on memory and concentration. We wished to investigate if individual subject hypometabolic regions are stable over time in longitudinal follow up imaging. We hypothesize that the downstream consequences of mTBI that may lead to neurodegenerative processes could be due to persistent regional hypometabolism. Positron emission tomography with [18F]-fluorodeoxyglucose (FDG-PET) imaging of cerebral glucose metabolism was used to assess brain metabolism in 13 veterans (mTBI) with mean age 31.7 ± 8.0 (range 24-49) years. Mean time between first scan and second scan was 3.73 ± 0.8 (range 2.2-4.7) years. We performed a longitudinal comparison with the follow up FDG-PET scans to determine if the hypometabolic differences altered as compared to FDG-PET from a group of age-matched community controls (n=9). Analysis included anatomic standardization through linear scaling/non-linear warping to the brain atlas followed by normalization to a mean whole brain metabolism. We used an automated program to segment the brain into 19 independent anatomical regions. The results indicated that metabolism in 3 regions in mTBI subjects were more than 2 standard deviations below the mean of controls: (9/13 right frontal, 9/13 right medial frontal, and 6/13 left medial frontal and these regions then resolved to within the normal range by scan 2 in 8/9, 7/9 and 5/6 subjects (78-89%)). We also found one region, left frontal, that was hypometabolic in 3 subjects and became further decreased by scan 2 in 2/3 mTBI (67%). These results indicate that hypometabolic regions in veterans with multiple mTBI may be normalizing slowly over time. This could be in part due to compensatory mechanisms related to a functional metabolic reserve in the brains of these subjects.

While increased female labor force participation has many positive implications, it may also have unintended negative consequences for women, such an increased risk of experiencing domestic violence. Our research aims to see whether there is a positive correlation between women’s labor force participation and the physical or sexual violence they face in the household, using wage, industry, and violence data collected from twenty-four regions in Colombia over eight years. We began by constructing a measure of random variation in labor demand to ensure that it was unrelated to changes in regional labor supply, thereby reducing potential endogeneity problems. Next we constructed two fixed effects regression models that estimated the relationship between work and each type of domestic violence. We found that there is a positive correlation between work and physical violence, but only among women with less education. We then altered our models to test the possibility that other measures of a woman’s initial level of bargaining power affects her likelihood of experiencing sexual violence upon entering the labor force. We are currently testing various violence theories that could explain why some working women are unable to leverage their increased intra-household bargaining power in order to reduce the violence they face. Ultimately, our findings have important implications for policymakers interested in women empowerment. In areas that experience job expansions for women, particularly those jobs that hire women with low education, policymakers should consider providing resources and programs that help reduce the increased domestic violence women may face.

Polyploidy, or an increase in whole sets of chromosomes, can result from a whole genome duplication event within an individual species (autopolyploidy) or following a hybridization event between two species (allopolyploidy). Determining the number and origin of genomes present within an individual will allow us to identify genomic mechanisms behind the emergence of plant diversity. Using a phylogenetic approach, our goal is to estimate the relative frequencies of the two mechanisms of polyploidy and their relationships to the diversity of flowers and species within the plant genus Thalictrum (meadow-rues), which contains numerous shifts in ploidy number, ranging from diploid (2x) to 24x. Since chloroplast DNA and nuclear ribosomal regions tend to underestimate the role of hybridization, I am using single-copy nuclear genes to uncover potential hybridization signals leading to allopolyploidy. This information will be used to compare the relative numbers of autopolyploids and allopolyploids in the genus, and to estimate ancestors of allopolyploid species. The data emerging from this analysis lead us to propose that major polyploid groups radiated after autopolyploidy events, while allopolyploidy contributed to further speciation events within these groups. The single-copy nuclear gene analysis also suggests that there may have been hybridization between the two clades containing dioecious species. We propose that hybridization between polyploids, biogeographical proximity, and the emergence of specialized floral morphologies, such as separate sexes, may be connected by a reticulated history. Analysis of additional single-copy nuclear genes and an updated phylogeny will be used to support these conclusions in future studies. My research project will test the relationships between the number and origin of genomes present within an individual and the diversity of species and flowers. The conclusions from my project allow us to identify the genomic mechanisms behind the emergence of plant diversity.

An automated wave plate rotator (AWPR) is an optical device that allows a user to remotely and precisely control the polarization of a laser beam. AWPR's are used in optical research studies of materials by providing optical intensity or polarization control. Commercial AWPR's are expensive and here I present the design and implementation of a low-cost alternative that utilizes a programmable microcontroller, stepper motor, and a few electronic components. The AWPR will be remotely controlled via Matlab and will include features such as home detection and position feedback. Results on position precision and accuracy will be reported.

There is theoretical and empirical support for normalization in human vision, the idea that the human visual system adapts, over the long-term, to chromatic regularities in the environment. Here, we analyze images of natural scenes to determine whether normalization can occur independently and differently for bright and dark colors. This goal is motivated by psychophysical evidence of regular differences in chromaticity between contextually bright versus dark colored stimuli. Finding regular differences in chromaticity between bright and dark portions of scenes, to which the visual system might adapt in the long-term, would link psychophysical results to the larger body of normalization theory. The present analysis models human long- (L), middle- (M), and short- (S) wavelength-sensitive cone photoreceptor excitations pixel-by-pixel for each image from numerous vision science databases. We analyze the relation of these cone excitations to variation in light level, using different sampling strategies, including variation in size and physiological relevance of samples. Using physiologically inspired models of chromaticity to sample images at different spatial scales allows us to explore the dual roles of local spatial and chromatic contrast in normalization, as well as the efficacy and consequences of such models. Our analysis is meant to highlight important trends in natural images to which the visual system might be normalizing. Describing chromatic differences by light level in a variety of scenes will allow us to define categories of images by content, which will provide a theoretical framework to explain our psychophysical results in terms of normalization. Finding a regular shift in chromaticity between high and low light levels, across models and within images of a given category, would suggest that observed psychophysical shifts in red-green balance between bright and dark colors are a product of long term adaptation to the natural environment.

One of the primary threats to agriculture is high salinity in soil, caused by the evaporation of water, which leaves behind dissolved ions that accumulate over time and pose a threat to non-salt-tolerant crops. The United Nations estimates that high salinity renders about half of the world’s agricultural land unusable, limiting the food available to feed our growing population. We are investigating salt-response mechanisms in Arabidopsis thaliana, a popular model organism for plants. A. thaliana has been found to share some of the genes involved in the salt-stress response of numerous crops, including maize and tomatoes. Our study examines HAC1, which is known to function as a histone acetyl transferase (HAT) and can bind transcription factors. HATs have previously been shown to bind DNA promoter elements of AtBZIP17, a membrane-associated transcription factor that is normally located in the endoplasmic reticulum. Our additional protein of interest, AtHB7 is known to be downstream of the transcription factor AtBZIP17 in the salt-stress pathway. AtHB7 is known to act in a signal transduction pathway as a negative regulator of growth in response to drought or salt-stress. AtHAC1 has been shown to be up-regulated as part of the salt-stress response. We propose that within the salt-stress response of Arabidopsis, AtHAC1 goes on to up-regulate the expression of AtHB7 by acetylating its promoters. We are addressing this hypothesis by examining the gene expression of AtHAC1 and AtHB7 of Arabidopsis root tissue in wild-type and hac1 mutants exposed to varying concentrations and durations of salt stress. We are unable to discern a direct relation between functioning AtHAC1 and expression levels of AtHB7, but we have identified AtBZIP17 as a future candidate for examining gene expression levels as related to AtHAC1 and AtHB7.

Telomeres are repetitive DNA sequences located at the extremities of linear chromosomes. With each cell division, telomeres shorten due to the inability of DNA polymerase to complete DNA replication at the 3’-end of the template strand. Telomere shortening occurs in primary cells during aging and in laboratory cultured cells with passage. It has also been observed in patients who receive myeloablative conditioning and bone marrow transplant(BMT) for certain blood cancers, which is likely due to a burst of cell division in the newly engrafted bone marrow stem cells as they reconstitute normal blood cell production known as hematopoiesis. In hematopoietic stem cell gene therapy, the contribution of the gene-modified pool to hematopoiesis can be increased by including the chemotherapy resistance gene, methylguanine methyltransferase P140K(MGMT^P140K), in the gene therapy vector. The addition of a chemotherapy resistance gene gives a selective growth advantage to the gene-modified cells following a low dose chemotherapy regimen of O6-benzylguanine(O6BG) and bis-chloroethylnitrosourea(BCNU). Due to the subsequent selective growth advantage of MGMTP140Kgene-modified cells, we hypothesized that this clinical protocol may cause further acceleration of telomere shortening in peripheral blood leukocytes. To test this, we used a quantitative PCR approach to perform longitudinal assessment of telomeres in 5 animals. This assay suggested that in animals with polyclonal engraftment of MGMT^P140Kcells, telomere length is unaffected following BMT and rounds of chemo-selection. In contrast, one animal with a dominant blood cell clone with integration in the TRIM44 homolog displayed drastic shortening in telomere length. It is still unknown whether this clone began with a shorter telomere or experienced telomere shortening as a result of rapid proliferation. Despite a decrease in telomere length, no clinical adverse events were observed. This qPCR assay has provided supplementary support to validate the safety and efficacy of the MGMT^P140Kchemoselection approach specifically in animals with polyclonal reconstitution.

A number of neuropsychiatric disorders including depression, autistim spectrum disorders, Parkinson’s disease, bi-polar disorder etc. are characterized by deficits in the ability to make decisions under changing reward contingencies. This ability to change responses when outcomes change is an executive function commonly termed cognitive flexibility. Current research has yet to yield a clear understanding of how these various diseases result in this common deficit at the neurobiological level. A common task used to test cognitive flexibility across species is known as reversal learning. Previous studies have shown that manipulations of serotonin (5-HT) and dopamine (DA) affect cognitive flexibility in tasks such as reversal learning. Importantly, these two neurotransmitters are known to play a role in a variety of neuropsychiatric conditions, including the ones mentioned, raising the possibility that a common mechanism may play a role across diseases. The lateral habenula (LHb) may be a key structure in mediating reversal learning as it is known to affect transmission of 5-HT and DA. Behaviorally, the LHb is thought to provide an error signal during decision making tasks making it likely that it is critically involved in tasks requiring learning from errors such as reversal learning. To test this hypothesis, a maze based probabilistic reversal learning task (PRL) with male long-evans rats was used to examine the role of the LHb via neurotransmitter inactivation with the gamma-aminobutyric acid (GABA) agonists baclofen and muscimol. Results reveal that inactivation of the LHb causes fewer overall reversals than rats injected with a saline control. Additionally, inactivation led to more errors per reversal than saline. Further analysis of error measures will reveal additional mechanisms, e.g. sensitivity to positive or negative feedback that underlies impaired performance during inactivation. These findings suggest that the LHb is important for learning and/or implementing beneficial behaviors when reward contingencies change.

TAF1 and TAF7 are two protein subunits of the general transcription factor complex TFIID, which comprises part of a larger protein complex essential in regulation of gene transcription and ultimately cell division. TAF7 binding to TAF1 inhibits TAF1’s histone acetyltransferase activity, which is required for cell proliferation. Interactions between TAF1 and TAF7 are influenced by TAF7’s phosphorylation state. In cancers, the cell cycle often proceeds without the same regulation mechanisms present in normal cells, which may contribute to the uncontrolled cell division observed in tumors. Therefore, we are interested in whether the abundance of TAF7 in its phosphorylated and nonphosphorylated states differs in normal and cancer cell lines in a manner which may implicate TAF7 in the proliferation of cancer. I have characterized an antibody, anti-pS264, that specifically recognizes TAF7 in its phosphorylated state. The experiments involved analyzing purified TAF7 and whole cell lysates by SDS-polyacrylamide gel electrophoresis and western blotting. I found that anti-pS264 recognizes TAF7 expressed in insect and mammalian cells but not in E. coli, which lacks enzymes capable of phosphorylating TAF7. Lysates treated with phosphatase enzymes, which dephosphorylate proteins, showed no protein recognition by anti-pS264. Subsequent addition of phosphatase inhibitors yielded reappearance of the TAF7 band, further verifying that the antibody recognizes TAF7 only in its phosphorylated form. I have begun screening TAF7 phosphorylation levels in a number of normal and cancer cell lines and in cell cultures synchronized at different stages of the cell cycle. Early results suggest that TAF7 expression and phosphorylation levels vary between cancer and normal cell lines. Varying amounts of phosphorylated and nonphosphorylated TAF7 observed in different cell types and under different conditions will give insight into the role of TAF7 and TAF1 in the regulation of cell division in cancer and, ultimately, into potential cancer therapy applications.

Pesticides are commonly used as crop protection products. While they aid crop growth by killing weeds and insects that are harmful to the plants, they could also have detrimental effects on many others, including microbes, animals and humans. The 16s ribosomal RNA project conducted at the Faustman lab investigates the effect of pesticide exposure on the microorganisms that typically inhabit an individual’s mouth. Through identifying the individual’s oral microbiome composition, not only are we hoping to understand how exposure to pesticides modifies the oral microbiome, but we also expect to find bacteria that are consistent with exposure to farming. The buccal swabs were collected from adults and children, who live and work in the Yakima Valley on apple farms. About 25% of these individuals were exposed to organophosphate pesticides, as detected by urine and blood samples that were collected alongside the buccal samples. The buccal samples of each individual will be amplified using targeted 16s ribosomal polymerase chain reaction (PCR). The amplified RNA will then be sequenced by the Ion Torrent to determine the phylogenetic composition of the microbiome in buccal samples. These analyses will help us find the differences in microbiome composition and metabolic patterns between samples collected from different agricultural seasons (pesticide use and non-spray seasons), and between children and adult samples. This year we will be advancing from last year’s 16s ribosomal RNA research progress in sequencing for taxonomic identification. If a correlation among the host genotypes, oral microbiome and pesticide exposure is found it would allow us to assess risk differences and define metabolic biomarkers more accurately for exposed populations.

Arenediazonium salt modification (also known as diazotization) is a useful and highly industrializable reaction, providing an intermediary for functionalization of aromatics into aryl- compounds, phenols and azo dyes. Graphene, a 2-dimensional, mechanically strong, electrically conductive, latticed aromatic, has chemical properties similar to formally defined aromatic compounds, due to the sp2 hybridization of the carbon atoms. Modification of graphene by diazotization could provide an efficient, if not elegant, mechanism for functionalization of graphene. Should the method produce a moderate yield, most functional groups can be added to graphene via diazo-salt replacement or conversion. The coupling of a diazo-graphene compound with phenol groups to form diazo-dye graphene (DDG) will prove the diazotization mechanism works. The tunability of DDG’s surface geometry provides an attractive avenue for photovoltaic research, particularly applicable to energy storage, or photo-optics. Fourier Transform Infrared Spectra (FT-IR) with the KBr method, as well as Raman spectroscopy will be used to monitor the presence of functional groups through each step of the mechanism. Atomic force microscopy (AFM), in conjunction with UV exposure, will be used to confirm geometric isomerism of DDG and the interaction with light. There is a large scope for the functionalization of graphene through this well-established organic method, which can ultimately be expanded upon with other carbon nanotubes or particles, for many future applications.

Associations between location and reward are responsible for behaviors essential to an animal’s survival such as foraging. Place cells in the hippocampus may play a role since they fire in response to an animal entering a specific region of space denoted as the cell’s “place field”. Place cells are also notable for exhibiting phase precession in which firing occurs at earlier phases of the theta component of surrounding neural oscillations as the animal traverses the place field. Our study examined dynamic properties of this location-reward network by recording hippocampal place cells from 4 rats trained on a probability-discounting task. In this spatial decision making task, rats traversed a circular track in which the animal chose between receiving a small, ‘certain’ reward or large ‘risky’ reward that is delivered with varying probability, depending on which of two doors the animal passed through. Each daily session contained blocks of trials: the probability of receiving the large reward was reduced across blocks, while the certain door remained associated with a 100% probability of receiving the small reward. Each probability block began with 8 forced-choice trials in which only a single door (4 risky and 4 certain) was open. This was followed by 8 free choice trials in which the 2 doors were open simultaneously. Preliminary analysis provides strong evidence that place fields differ (or ‘remap’) between forced and free-choice trials, and also across probability and trial outcomes. Further, theta power in the hippocampal local field potential was modulated by probability and outcome at locations associated with salient events. Phase precession will be investigated as a feature of place cell firing that is likely to change in correlation with the remapping and theta modulation described above. Thus, place fields depend not only on spatial features, but also risk and agency (degree of free choice).

Growing prevalence of age-related neurodegenerative disorders has precipitated the need for

sensitive early indicators of disease allowing targeted, early intervention. Alzheimer’s disease

(AD), the most common form of dementia in the United States, is reliably predicted by

amnestic mild cognitive impairment (aMCI) as revealed by cognitive testing; however,

cognitive impairment likely follows substantial brain changes including changes in the

structure and function of the hippocampus. Preliminary evidence indicates that hippocampal

function can be indexed by the novelty preference effect, a measure of recognition memory

which describes the tendency to look longer at novel images than familiar ones. This study

will investigate whether eye movements during novelty preference tasks can help identify

individuals at risk for aMCI and AD. We collected data from 54 participants (n=29 women)

aged 65 and above (mean=78.6 years) recruited from Group Health Cooperative’s Adult

Changes in Thought study. All participants completed cognitive testing with a standard

neuropsychological battery and a Magnetic Resonance Imaging (MRI) session with

structural, diffusion, and resting state functional scans. During structural scanning, we

tracked participants’ eye movements while they viewed a series of images that were either

repeated, or otherwise completely or partially novel. We are in the midst of analyzing data

from the scans, and will assess the recognition memory of each individual by calculating the

proportion of time they spend looking at novel relative to familiar stimuli. We will then

determine whether a decreased tendency to look longer at novel images is correlated with

poor cognitive test performance, atrophy of the hippocampus and other memory-related brain

regions, and structural and functional connections between these regions. If novelty

preference is related to cognitive status and other markers of neurodegenerative disease, these

eye tracking tasks may be a practical and cost-effective way to help identify individuals at

risk for later dementia.

The purpose of this project is to investigate the optic disc perfusion differences in glaucomatous eyes and normal eyes using Cirrus 5000 HD-OCT based optical microangiography (OMAG). Eyes of normal subjects and glaucoma patients were scanned with a 67 kHz Cirrus 5000 HD-OCT based OMAG prototype system (Zeiss, Dublin, CA; 245x245 raster cube scan pattern over a 2.4x2.4mm2 area centered at the optic nerve head (ONH) region). OMAG enface images for pre-lamina layer, LC, and the entire ONH (from ILM to the outer boundary of choroid) were generated using maximum projection. The optic disc margin was defined using Bruch’s membrane opening and was manually delineated based on the structural enface image. Optic disc perfusion was measured by calculating the mean flow intensity within the optic disc region, and compared between glaucomatous and normal eyes. Repeated measures analysis were performed to compare optic disc perfusion between glaucoma and normal eyes and accounted for cluster effect of including both eyes from the same subject. P<0.05 was considered as statistically significant. 25 eyes from 13 glaucoma subjects (mean visual field mean deviation -4.43±5.19 dB) and 15 eyes from 8 age-matched normal subjects were recruited. Optic disc perfusion was statistically significantly lower in glaucomatous eyes compared to normal eyes in the entire ONH (0.30±0.03 vs 0.27±0.03, p=0.022). Optic disc perfusion detected by OMAG was significantly lower in glaucomatous eyes than normal controls. Optic disc perfusion measurement may enable detection and monitoring of glaucoma.

A disulfide bond is a covalent bond that crosslinks sulfur atoms between two cysteine amino acids. Disulfides are an important part of protein structure, and they generally increase a protein’s stability. They are often found in proteins secreted to the extracellular medium, such as antibodies or hormones. Disulfides are unable to form in the reducing environment of the cytoplasm; this makes the expression and purification of proteins that contain them difficult. Recently, it was discovered that the natively secreted Escherichia coli protein OsmY (osmotically inducible protein Y) could be hijacked to allow for secretion of recombinant proteins containing disulfide bonds to the extracellular environment by genetic fusion. E. coli is a Gram-negative bacterium, thus it possesses two membranes surrounding the cytosol, with a periplasmic space in between. Export of OsmY from the cytosol to the periplasm occurs via the conserved Sec machinery. However, the mechanism responsible for export of OsmY across the outer membrane is currently unknown. Our investigation is aimed at gaining a better understanding of the mechanism responsible for OsmY’s outer membrane translocation. The mature OsmY protein consists of two domains: an unstructured N-terminal domain, and a BON (bacterial OsmY and nodulation) domain, which has putative membrane binding activity. We are assessing the role of each domain on outer membrane translocation independently using recombinant expression constructs and cellular localization assays. Our goal is to find the minimum segment of OsmY that is sufficient to mediate extracellular localization. Our research could allow for generation of improved secretion carrier constructs, which in turn could enhance production of protein reagents and therapeutics.

HIV-1 binds to cells using a combination of specific receptors on T-cells: the CD4 receptor (primarily) and the CCR5 receptor (during early stages of the disease). Individuals with a homozygous CCR5-delta-32 mutation, a 32 base pair deletion in the CCR5 gene are resistant to HIV-1. A patient that received a transplantation of hematopoietic stem cells (HSC) containing the CCR5-delta 32 mutation was cured from AIDS. Using this information, we aim to induce CCR5 gene knockout using Zinc-Finger Nucleases (ZFNs) delivered by a helper-dependent (HD) adenovirus vector (HD-AD5/35-ZFN), an alternative to currently employed delivery systems including electroporation with mRNA. The ZFN delivered by the vector introduces double-strand DNA breaks in CCR5 gene. These breaks are then repaired by a cell mechanism called non-homologous end-joining (NHEJ), which usually produces mutations disrupting the reading frame. We conducted preliminary experiments using HeLa-TZM-bl cell line that expresses both the CD4 and CCR5 receptors and infected them with HD-Ad5/35-ZFN. Upon infection with the HD-Ad5/35-ZFN vectors, the cells were analyzed by flow cytometry in order to measure the amount of CCR5 receptors on the cell surface. As the concentration of the virus added to the cells increased, the amount of CCR5 receptors decreased. This proves the link between the activity of the ZFN and the percentage of cells with the receptor in a specific subset of the population. Along with the CCR5 disruption at the protein level we examined the knockout at the genomic level using a mismatch sensitive PCR assay. Current data shows us that that HD-Ad5/35-ZFN is an effective vector for the delivery of ZFNs to cells. Future work will focus on the optimization of CCR5 knockout in HSCs and transplantation of the modified cells into mice.

The linguistic word, defined as the smallest unit of language that can be used in isolation, was not introduced to Chinese culture until Western influence in the 20th century. Instead, Mandarin Chinese is comprised of individual characters that each represents a morpheme, which is the smallest unit of language that contains meaning. Morphemes in English include root words, prefixes, and suffixes. In Chinese, there are multiple types of compounds comprised of at least two morphemes that have come to embody a specific meaning when used together. The goal of this study is to investigate whether any of these compounds may now be considered a word by native Chinese speakers with no formal linguistic training. To do so, this experiment examines event-related potentials, which are averaged brain waves recorded from locations across the scalp, from native Chinese speakers while they read Chinese sentences. Some trials involve situations where the second morpheme of a compound is removed mid-sentence. Two types of compounds are manipulated: true compounds, where each morpheme contributes meaning to the overall compound, and core compounds, where one morpheme contains the central meaning. It is hypothesized that leaving off the second morpheme of true compounds will result in a typical semantic error that produces a negative wave in the subject’s brain approximately 400 seconds after the error in meaning. On the other hand, the unique nature of core compounds, where the second morpheme is often semantically vacuous yet necessary for completion purposes, allows for a range of possible responses. As such, this study primarily focuses on comparing the brain responses to ill-formed core compounds to the brain responses to ill-formed true compounds. Depending on the results, this study may offer insight on whether native Chinese speakers have come to internalize the Western concept of a word within their own language.

TFIID is a general transcription factor that is part of the transcription preinitiation complex. TAF1 is a subunit of TFIID and it binds to DNA at the promoter region of many genes. Mutations in the protein sequence of TAF1 or other TFIID proteins could cause transcription problems which can lead to diseases ranging from neurological disorders to anemia and even cancer. Previous research has shown that when TAF7 binds to TAF1 transcription is repressed. We want to figure out if the 250 amino acids on the C-terminal of TAF7 binds to TAF1 in the absence of a known N-terminal binding domain. We also want to know if phosphorylation regulates binding, so we mutated serine 264, an amino acid that is commonly phosphorylated in TAF7, to alanine (A) which cannot be phosphorylated or aspartic acid (D) which mimics a phosphorylated amino acid. In my work, I learned to create recombinant insect viruses to express specific parts of TAF1, TAF2, and TAF7, in the wild-type form or with specific amino acids mutated. The next step is to analyze the lysates of the infected insect cells by western blotting. We have labeled the TAFs with specific tags so we can do affinity purification using beads specific for the tag to pull the protein down. From the western blot, we can see if TAF1 was pulled down with TAF7, and vice versa, and we can determine what part of TAF7 is then responsible for this binding pattern. This can tell us more about what specific parts of TAF7 are important in binding TAF1, and thus we will learn more about which mutations could be potentially harmful and contributing to human diseases.

Perfluorinated compounds (PFCs) have been manufactured for over 55 years. Many industrial and consumer products worldwide have been made from these compounds. Perfluorooctanoic acid (PFOA) and Perfluorooctanesulfonic acid (PFOS) are perfluorinated hydrocarbons that can be used as a repellent of water; so they are used to resist stains for clothes, carpets, food containers, floor polishes and sometimes used in insecticide formulations. The environmental concerns for these compounds arise due to their persistence, widespread detection, and potential to bioaccumulate. They also have been detected in human blood and breast milk. Human exposure to these contaminants could be due to ingestion of contaminated foods, including drinking water and inhalation of dust. This study was performed to determine the concentration of PFCs, especially the PFOA and the PFOS in dust samples from Vancouver, Washington, USA. PFCs were extracted from the dust using acetonitrile and sonication and analyzed by liquid chromatography-tandem mass spectrometry (LCMSMS). The average concentration of PFOA and PFOS detected were 109ng/g and 78ng/g with the range from 66 to 223ng/g and 43 to 155ng/g respectively. These averages of PFCs were much lower than the findings in 2008 of household dust samples from Ohio and North Carolina USA.

Is there a correlation between hormonal contraceptive intake during the peri-conception period and adverse birth outcomes, such as birth defects and fetal death? In 1990, Braken concluded that there was no correlation between hormonal contraceptive intake and adverse birth outcomes. Although methods of contraception have changed, we also hypothesis that there is no correlation between contraceptive intake and negative birth outcomes. We are using a systematic review methodology according to the United States Preventive Services Task Force. This research is being performed in conjunction with Centers for Disease Control and Prevention. We performed a systematic search of articles through PUBmed and Embase. We systematically abstracted data using a validated form of Agency for Healthcare Research and Quality. To grade the included articles we are using a quality framework to categorize articles into different levels of quality based on selection bias, information bias, confounding factors, and precision. Potential results from this systematic review may suggest a strong or weak correlation between contraceptive intake and negative birth outcomes or an absence of an association, in general. If we are able, we will be performing a meta-analysis of the data. Through this analysis, we hope to ensure the safety of contraceptives so that its use may become more prevalent. We anticipate further research for newer methods of contraception.

Major challenges in tissue engineering come from difficulties inherent to the process of generating suitable biocompatible scaffolds that are needed to promote desired cell formation and growth. Several of these difficulties in controlling the spatial, temporal, mechanical and biochemical architectures could be overcome by taking advantage of the recent developments that make high-resolution stereolithography (3D printing) technology fast and affordable. This project focuses on the development of protocol for the formation of photo-induced carboxybetaine acrylamide (CBAA) hydrogels using an extrusion-based 3D printer for rapid prototyping of biocompatible tissue scaffolds. CBAA hydrogel formation using photolithography was investigated by considering multiple aspects relevant to their formation. Multiple photocatalysts and crosslinking agents were considered for the strength and localization of activity. The effects of extrusion rate and ultraviolet wavelength intensity were also considered. Then the temporal viability of cells on these hydrogel environments was evaluated. A key challenge affecting hydrogel formation persists in the solvation of the photocatalyst into the aqueous CBAA monomer solution. Further, specificity of reaction at the desired location remains a challenge. That being the case, the ability of cellular proliferation to occur shows potential for improvement. Results indicate that photolithographic techniques used for the rapid prototyping of biocompatible CBAA hydrogels has the potential to address many of the difficulties encountered in the field of tissue engineering. Further investigation into alternative photocatalysts and using a higher resolution 3D printer may prolong cell viability.

The collective and coherent excitations of conduction electrons at the surface of metallic nanoparticles (NPs) are known as localized surface plasmon resonances (LSPRs, or surface plasmons), and are caused by the absorption of light incident on the surfaces of the NPs. These LSPRs can decay radiatively (by emitting light) or non-radiatively, such as by generating heat. It has been found that in certain NP systems of the right composition and morphology, the temperature profile of the system can be spatially tuned so as to cause significant temperature differences even within the same diffraction-limited spot in the steady state. This is unusual in that thermal diffusion tends to cause a uniform temperature change throughout systems of much higher thermal conductivity than their environment, regardless of the spatial location at which the heat power is supplied. However, in some systems which support a type of resonance phenomenon known as Fano resonance, the phenomenon of Fano resonance can be utilized to spatially direct the locations at which heat power, as a function of the locally scattered electric field caused by LSPRs, is supplied. Further, the Fano resonance ensures that the resulting temperature increases in the system correspond to these locations in the steady-state. This ability to localize heat in nanoscale domains only by shining light on the system has far-reaching implications for applications such as chemical catalysis, thermal cloaking, and biological thermal activation. In this presentation we illustrate, using a coupled dipole model, the mechanism by which the electromagnetic near-field strength can be spatially tuned. We also showcase several systems of metal NPs which allow such temperature manipulation, and discuss possible applications. 

Bumblebees pollinate tomatoes and many other food plants, providing great economic value; unfortunately, declines in multiple bumblebee species across North America have been documented.  Commercial greenhouse tomato pollination is often done by machine-"buzzing" tomato flowers to increase self-pollination, but a pilot study found that this treatment produced smaller fruit than bumblebee pollination.  We asked:  how important is cross-pollination by bumblebees to fruit production and to plant reproductive success?  We treated 24 tomato plants (Solanum lycopersicum var.) with two self-pollination methods (natural selfing and hand-"buzzed" selfing) and two cross-pollination methods (bumblebee crossing and hand-"buzzed" crossing).  We measured volume and seed number per fruit, as well as percent germination of seeds produced form each treatment.  Percent germination was highest in the hand-"buzzed" selfing treatment, suggesting that bees may introduce variables that reduce seed viability or that there is a fitness trade-off between seed number and total germination.  Nevertheless, we found that cross-pollination by bumblebees resulted in the highest volume and greatest number of seeds per fruit over all other methods of pollination.  These results provide clear support for the value of the ecosystem services bumblebees provide.  Conservation and agricultural practices should be further considered to promote healthy bumblebee populations. 

Light Chain Amyloidosis is a disease defined by the deposition of aggregated immunoglobulin light chains in the form of fibrils in several organs. While these fibers are insoluble, the soluble oligomers that precede them in the pathway involving unfolding and then aggregation are toxic to the cell. A novel secondary structure, alpha sheet, was hypothesized to be the cause of toxicity. This structure is rarely seen in normal proteins, but it is populated in all peptides and proteins involved in amyloid diseases studied so far in the lab. In addition, the correlation between thermal stability and fibril formation has led to the study of the impact of amino acid mutations on the unfolding pathway. Unfolding and subsequent misfolding can increase the chances of fibril formation. As is the case for most amyloid diseases, Light Chain Amyloidosis is fatal with no known cure. The determination of specific patterns and mutations that destabilize the protein and lead to increased toxicity is important to better understand how to intervene. In order to identify these locations, we are comparing two light chains, one non-amyloidogenic and the other amyloidogenic. By analyzing their differences, we hope to determine areas that contribute to aggregation. We performed molecular dynamics simulations of each protein and analyzed their unfolding pathways. Areas of interest are the complementary determining regions, the variable regions in light chains responsible for antigen binding, as they are more prone to amino acid mutations. Specific residues' interactions have also been identified that are integral components to stabilizing the folded state. The dissolution of these interactions leads to fluctuations within the protein and subsequent unfolding. A detailed molecular characterization of these conformational changes should aid in drug design.

Alzheimer’s disease (AD) is an age-related neurodegenerative disorder afflicting millions of people worldwide. Biomarker research seeks to identify molecules to diagnose, track, and differentiate AD from similar diseases like Parkinson’s disease (PD). For instance biomarker candidate alpha synuclein (α-syn) is higher in cerebrospinal fluid (CSF) from AD patients and lower in PD. However, CSF collection involves an invasive procedure, while blood, though less invasive to collect, is affected by peripheral systems. This has led researchers to consider a more accessible fluid, saliva, as a source for biomarkers. However decreased salivary production related to age, compounded by dry mouth in AD patients, can complicate collection of an adequate amount of saliva via passive donation. To facilitate collection, some studies have used materials such as cotton to stimulate saliva production; however, the materials used may affect native protein concentration. For instance α-syn is known to stick to cotton, artificially lowering the apparent concentration of this protein in samples collected using cotton stimulation. Thus a systematic examination of the effect of various materials on sample composition is needed to determine ideal conditions for stimulated saliva collection. In this study I sought to determine how common materials impact salivary α-syn and total protein concentrations in vitro. Ten materials were incubated in pooled reference saliva samples for 30 minutes and 17.5 hours. α-syn was quantified by Luminex while total protein was quantified by a Bradford assay. Cotton (A, 30min and 17.5hr) and gum (B, 30min) significantly reduced α-syn, while harder surfaced materials generally did not have a significant effect. Total protein was not significantly influenced by the tested materials. The results of this experiment demonstrate that several materials appear to have no impact on either total protein or α-syn concentration, and may be suitable for increasing sample availability without drastically altering native protein composition.

What is the effect of sensory deprivation on brain development? Ocular Dominance Columns (ODCs) have been extensively used to study the mechanism of cortical plasticity, although the specific function of ODCs has remained elusive. Using a combination of transneuronal tracing, in situ hybridization for the immediate early gene Zif268 and electrophysiological recordings, our lab recently showed that the primary visual cortex (V1) in pigmented rats has ODCs, and these ODCs correlate with callosal inputs from the opposite hemispheres. Using similar methods, my project aims to understand the effect of monocular deprivation on the newly discovered system of ODCs in rat visual cortex. I hypothesized that the manipulation of visual input will impact the development and recovery of eye-specific circuitry. Since the distribution of callosal connections is closely associated with ODCs, I am also interested in the effect of monocular deprivation on the callosal pattern. By disrupting the visual input permanently or temporarily in early postnatal life, our lab has been able to measure the deleterious effects on map development, critical periods for these effects and potential for recovery. My project uses monocular eyelid suture to study the effect of monocular deprivation on the organization of ODCs and callosal connections in pigmented rats. By using our experience-dependent model we expect to elucidate the principles and mechanisms underlying the development of cortical modular architecture in mammals. If our hypothesis is supported, our novel model may shed new lights on plasticity research, which can guide future clinical studies on the treatment and prognosis of patients suffering eye defects during early development by either innate or incidental pathological causes.

Members of the genus Passiflora, commonly known as passion flowers, have the most morphologically diverse leaves of any plant genus. For a generation, scientists have hypothesized that this incredible leaf diversity may be the result of a complicated co-evolution with the animals that herbivorize them, the Heliconiinae butterflies. As Heliconiinae caterpillars evolved to prey exclusively on Passiflora leaves, the genus has responded throughout evolutionary history with fascinating mimicry and a suite of toxins, in a particularly compelling example of co-evolution between a host and a specialized pest species. My project focuses on understanding how predation pressure by the butterflies may have led to the incredible leaf diversity in the Passiflora, using morphological analysis of leaf samples from the UW greenhouse collection. Beyond their toxicity, Passiflora leaves bear many other interesting markers that are part of their evolutionary history, such as size, coloration and leaf spots, which will be quantified in this project using photo-editing software tools to collect pixel counts and red/green/blue scale information. The morphological data will then be mapped onto the Passiflora phylogeny built by our lab and collaborators to ascertain any patterns that may exist in the morphological changes throughout this genus’ natural history. I predict that Passiflora that are preyed upon by a variety Heliconiinae species will display a higher number of egg spots than Passiflora that are preyed upon by fewer species, as caterpillars are cannibalistic and prefer not to lay eggs on colonized plants. I also predict that leaf shape and size will vary greatly throughout evolutionary history, as the more the leaves differ with time, the more difficult it is for Heliconiinae to hone in on a specific host to feed or lay eggs. The culmination of this data aims to illustrate the fascinating evolutionary history of Passiflora and its closely co-evolved herbivores.

Choroid, lying between retina and sclera, plays an important role in maintaining eye function and providing oxygen and nourishment to photoreceptors. Myopic eyes are thought to be modulated by the choroidal function. Many studies observed that the fluctuation of choroidal thickness (CT) during the day is evident and suggested that the fluctuation is shown to distract eye examination. However, none of them provided a comprehensive and precise study using a high resolution technique and they fail to keep the relation of CT to axial length (AL) of eye into consideration. Spectral Domain Optical Coherence Tomography (SD-OCT) used in our study is a non-contact imaging technology capturing detailed 3D image with a high speed image acquisition by reflecting light within optical scattering media. In this study, we hypothesized that the application of SD-OCT can provide us correlation between the diurnal fluctuation of CT and the AL of eye. To fulfill this goal, we use 140 kHZ SD-OCT to record the CT and AL of different subjects at three time points during one day. For its repeatability, one subject is chosen to perform the same measurement in five different days. By calibration of motion artifact and segmentation of different layers of human eye, the information of CT is generated and used to produce a 3D image for comparison. Also, the relation to AL and the fluctuation range is established after statistical analysis. We found that the variation of AL influences the variation of CT and the result after normalization of the two is different from single study of diurnal fluctuation of CT. In summary, if our finding of the correlation is universal, a better understanding of human eye structure and function is demonstrated, which will be beneficial in both clinical measurement and further analytical eye study.

Cancer cells can grow into life-threatening tumors and metastases partly because of their ability to evade the immune system. Antibody and cell-based immunotherapies are being developed to target this problem by boosting the immune response to cancer and delivering drugs directly to cancer cells. Immunotherapies are often more successful than standard treatments like chemotherapy because they are more-targeted, and have longer lasting effects and milder side-effects. T lymphocytes are a type of immune cell that kills pathogens, so they are a logical target for immunotherapies. Currently, bioengineers lack ligands that bind to T lymphocytes, which are needed to synthetically modify T lymphocytes for targeted drug delivery or immunomodulation. The purpose of this project is it to fill this gap and identify peptide-based ligands that bind to human CD28 and CD3e, which are surface proteins on T lymphocytes. To find peptide ligands, methods of phage display are used, which involves sequencing a sample from a large population of peptides that have been selected for binding to CD28 and CD3e. First, phage display and traditional Sanger sequencing were used to optimize the selection conditions and try to identify peptides that bind to CD28 and CD3e. However, due to the low throughput of Sanger sequencing, no peptide ligands were found. To overcome this challenge, we evaluated the use of high throughput Illumina Next Generation sequencing to identify target-binding peptide sequences. Next Generation sequencing is more effective than Sanger sequencing because it can quickly sequence millions of peptides and reveal false-positives due to preferentially amplified phage clones. Our analysis of Next Generation sequencing will be applied to find peptide ligands to address various diseases. Specifically, we will use Next Generation sequencing to find peptide ligands that bind to CD28 and CD3e and evaluate these peptides for their feasibility as cancer immunotherapy components.

According to the most universally used scale for measuring psychological distress, the Holms and Rahe Readjustment Scale, becoming a widow holds top rank for the most taxing life-event. Unless specifically defined otherwise, it is generally understood that the term ‘widow’ refers to a heterosexual female. Consequently, few studies highlight the psychological distress of self-identified lesbian widows, and even fewer examine resiliency. Resiliency is the capacity to recover from difficulties, where individuals use their personal skillsets to reengage themselves within their environment. To our knowledge, no current research directly compares features of both lesbian and heterosexual widow resiliency within the same qualitative analysis. We seek to understand key features and possible differences in resiliency between self-identified lesbian and self-identified heterosexual widows. Our intention is to learn what types of support best aid resiliency among widows, and whether these support systems are readily accessible to lesbian widows in mainstream heterosexist U.S. society. To measure resiliency, we are conducting voluntary, in person, one-on-one tape-recorded interviews of 15 lesbian and 15 heterosexual widows. Inclusion criteria required participants be at least 45 years of age and widowed for a minimum of one year at the time of interview. We analyzed interview data through the constant comparative analysis method. We here present a preliminary report on 6 widows from each group. We expect to find differences between groups in a variety of themes related to resiliency (i.e., perception of available support systems and/or effectives of support systems, if any.) Participants in the research will give us an understanding of the underlying attitudes and social dynamics that influence resiliency in this context, helping us identify the conceptual framework needed to best improve resiliency among lesbian and heterosexual widowed populations.

Previous studies show that females who have been sexually assaulted report increased alcohol use. In one study, PTSD symptoms were positively associated to coping-related drinking motives, however, PTSD was not associated with drinking for enhancement, conformity or social reasons. Another study found a positive relationship between motivation to drink for conformity and drinking behavior among women with PTSD. There is an inconsistency in the small amount of literature related to drinking motives of women with trauma. In addition, there is a lack of research focused on drinking motives among women with adult sexual abuse (ASA). We predicted that drinking motives would vary as ASA severity increased. A population of women (N=869) aged 21-35, were recruited from an urban community on the west coast. Participants were heavy episodic drinkers, single, and at risk of sexually transmitted infections. Four separate simple regression analyses examined the relationship between ASA severity and drinking motives (enhancement, coping, social, and conformity). Significant results suggested a positive relationship between ASA severity and two drinking motives: 1) drinking to conform and 2) drinking to cope. As ASA severity increased, participants were more likely to drink to cope and drink to conform. However, we found no significant relationship between ASA severity and drinking for social or enhancement reasons. Drinking alcohol is an unhealthy coping mechanism and has been associated with risk of re-victimization and participation in risky sexual behavior. A better understanding of drinking motives that increase alcohol consumption in women with ASA may help inform more accurate and effective interventions to lessen these risks to women.

Amyloid diseases have quickly become one of the most pressing health concerns facing our generation, encompassing such disorders as Alzheimer’s disease and type 2 diabetes. Prior molecular dynamics (MD) simulations suggest that these diseases share a common mechanism of toxicity, wherein misfolded proteins aggregate into soluble oligomers typified by novel α-sheet secondary structure. Peptides specifically designed to be complementary to these toxic species by adopting this backbone conformation effectively inhibit amyloidosis. In this study, we demonstrate the importance of sequence and residue chirality in determining aqueous solubility and inhibitor potency. We assembled nine structural isomers with identical sequence composition but variable sequence identity and chirality. Some of these peptides followed an alternating L/D amino acid template, which stabilizes α-sheet structure, while others had scrambled sequences. We then co-incubated the designs with Aβ42 (whose aggregation is associated with Alzheimer’s disease) and islet amyloid polypeptide precursor (type 2 diabetes) and monitored their aggregation. The hairpin designs incorporating alternating L/D amino acids were water-soluble and effectively inhibited amyloid formation, but the scrambled designs showed poor solubility and/or inhibitory properties. Interestingly, although our designs utilized the same twenty-three amino acids in all cases, their physical properties and chemical behaviors showed wide variation, affirming the importance of secondary structure (in particular α-sheet structure) in determining a peptide’s efficacy as an aggregation inhibitor and, ultimately, as a potential therapeutic agent. 

It has yet to be shown how endophytic bacteria are forming associations with their hosts in the natural environment. Endophytic associations have been shown to benefit several species of plants in a variety of ways including improved growth and increased drought tolerance. Utilizing these endophytic relationships has powerful implications for improving crop sustainability, alleviating pollutants in the environment through phytoremediation, and increasing the production of biofuels. Chemotaxis experiments were performed using known endophytes and a variety of sugars in order to determine the potential chemical signals which attract endophytes to host plants in the environment. Partially sealed microcapillary tubes were filled with different sugars and placed in a solution of known bacterial concentration over different time points. The bacteria were then able to move either into or away from the tube. After each time point, the contents of the microcapillary tube were plated and colonies were counted. This method of determining which chemical signals attract different endophyte species helps us to determine the mechanism by which endophytes locate and are able to associate with their hosts naturally in the environment. Chemotaxis results differered between endophyte species indicating that different chemicals may attract or repel different bacteria naturally in the environment. Further research should be done in order to clarify which chemicals have the strongest reations for different endophytes. 

Does the gender gap in college enrollment vary across states? Which localities encourage women and men to attend higher education institutions? This paper uses a multi-step, multivariate regression to gain insights into the relationships between female enrollment in higher education institutions, age at first marriage, political leaning of the state, and leadership positions held by women. In the 1960s, there were 1.60 males for every female graduating from a four-year college, compared to in 2003 where 1.33 females were graduating for every male in a four-year college in the U.S. In 1982, the number of females graduating from U.S. four-year colleges surpassed the number of graduating males. This paper explores several possible explanations for the reversal of the gender gap. While past work has focused on the role of secondary school performance and changes in the labor market composition, I also take into account the political affiliation of the state and changes in gender norms over time. The empirical framework uses state-level variation in these factors during the last two decades to better understand the increase in women's enrollment in insitutions of higher education. 

Professional sports are making great strides in their commitment to environmental sustainability. Currently, nearly 300 teams representing 20 different leagues and 14 countries are members of the Green Sports Alliance (GSA). The GSA is a non-profit organization that facilitates professional and collegiate sports teams in improving their environmental performance by offering information on sustainable programs and implementation incentives. This research explored the degree to which professional sports teams are improving their environmental performance. Specifically, this investigation assessed the 30 professional teams of the National Hockey League (NHL) using data provided in the 2014 NHL Sustainability Report, the first such report issued by any North American sports league or team. This report focused on venue and game day operations and examined areas such as carbon emissions, waste reduction, energy consumption, water use, and fan engagement. To evaluate environmental performance, the Global Reporting Initiative’s (GRI) G4 Sustainability Reporting Guidelines were used to assess, compare, and rank the teams from highest environmental performance to lowest environmental performance. The guidelines reviewed intent, metrics, and impact according to the information provided in the sustainability report. This research provided an informative and measurable profile of each team’s environmental performance. Unlike financial reporting, sustainability reporting is voluntary and there is no standardized procedure to assess environmental practices. By auditing the 2014 NHL Sustainability Report, this research elicited the ways in which transparency and metrics can be improved in sustainability reporting and how sustainability reporting can be implemented at various scales and in sectors beyond professional sports.

Previous research demonstrated via PET scans that crows differentiate between and remember faces of threatening vs. caring people, however still much is unknown about these complex creatures. The purpose of our study is to determine how the brain of a crow responds to the visual stimulation of seeing a dead crow vs. the auditory stimulation of hearing scolding calls elicited by other crows that view a dead crow. Molecular imaging with microPET and [F-18]FDG was used to investigate neural pathways underlying the crows’ responses. For stimulus condition, eyes-covered crows received 1mCi FDG ip. During uptake, awake crows were shown 1) a dead sparrow 2) a dead crow 3) scolding calls but no dead bird, in 1m on/30s off blocks for 11m. Blink rate during stimulation was recorded as a behavioral measure of interest. After stimulation, crows were anesthetized and imaged by microPET. Reconstructed images were aligned stereotactically to a crow atlas and structural MRI and globally normalized. For analysis, a longitudinal comparison was done to visualize the activated neural pathways. We used Neurostat/3D-SSP to view the statistical significance between the baseline (empty room with dead sparrow), auditory stimulus, and visual stimulus. The images were then normalized and compared statistically on a voxel-wise basis. The statistical algorithm performs an r-to-z transform and the statistical significance of the resultant z-score values is determined with a Type I error rate of p<0.05 (corresponding to Z > 4.0). The brain activity indicated there are no significant differences between responses to dead crows and response to the sounds of other crows viewing a dead crow. The activation is remarkably similar and has a bilateral activation in the HA, which may be responsible for arousal, and more specifically, arousal to danger.

p120 catenin is an essential protein in the development of embryos, it is associated with the regulation of cell migration and adhesion during development. We propose that p120 catenin phosphorylated on tyrosine residues binds to the cytoplasmic tail of E-cadherin to stabilize and strengthen adhesion. In contrast we propose that when p120 catenin is phosphorylated on serine or threonine residues, it is released into the cytosol to regulate Rho-GTPase to facilitate cell migration. Two regulatory domains of p120 catenin contains serine/threonine residues that can be phosphorylated to allow cell migration and adhesion. I am modifying the phosphorylation sites to amino acid residues that lack the ability to be phosphorylated (serine/threonine to alanine) or to amino acids that are constitutively charged to mimic phosphorylation (serine/threonine to glutamic acid). By modifying these residues, we hope to distinguish phosphorylation sites that are essential for the adhesion and migration of cells during development. To test these modified residues, we inject splice site antisense morpholino into one-celled zebrafish embryos to knockdown the endogenous expression of p120 catenin, thus inhibiting somite formation and dorsal axis extension. The modified p120 catenin mRNAs are co-injected with anti-sense morpholino and the development of the embryos are compared with the embryos that were rescued with wild-type p120 catenin mRNAs. Data from the injection of p120 catenin mRNAs with threonine 910 to alanine or glutamic acid mutation rescues normal phenotype. These data suggests that the phosphorylation of threonine 910 is not essential for cell motility. Currently, I am studying another phosphorylation sites on the C- terminal end of p120 catenin, serine 879 to see if it is essential for cell adhesion and migration.

Biofuel is nowadays a popular alternative to fossil fuel energy. Ethanol, by far the most common biofuel, can be made from sugars in lignocellulosic biomass. Moreover, lignocellulosic biomass can also be converted into valuable products, such as acetic acid, xylitol, and butanol. Therefore, raw biomass composition is an important factor influencing the final cost of biofuel production. The challenge of lignocellulosic biofuel production is the cost of the feedstock; the biomass cost makes up to 45% of operating costs in ethanol biorefinery. The objective of this project is to find a hybrid poplar clone with highest sugar content. High sugar content produces high ethanol yield, which results in a more economically feasible biofuel production process. The lignocellulosic biomass is composed of lignin, cellulose, and hemicellulose. The cellulose can be broken into monomeric glucose and hemicellulose into arabinose, xylose, galactose, glucose, and mannose that can be converted to ethanol and other products. In this project, lignin, sugars, ash, and extractive concentrations will be measured by Klason analysis, UV readings, and extraction process. After a series of experiments, the concentrations of sugars will be compared to select the hybrid poplar clone with highest sugar content. The clone that has the highest sugar content is hypothesized to be the most economically feasible feedstock for bioethanol production.

The goal of this research is to test and develop an aptamer labeled with zirconium-89 to target an Epidermal Growth Factor Receptor (EGFR) to allow in vivo PET imaging. Aptamers are novel binding reagents that are comprised of short strands of DNA or RNA. The aptamers we are using are DNA-based with synthetic side chains that afford optimal binding properties, as well as improved pharmacokinetics compared to RNA constructs. Our target, the EGFR, is overexpressed by many human cancers, including lung carcinomas, breast cancers, and gliomas, and is a prime target for developing novel targeting reagents to treat these tumors with more specificity. We have demonstrated specific EGFR binding in vitro through fluorescence microscopy with a fluorescently labeled version of our aptamer that bound significantly greater to EGFR positive cell lines than negative cells lines. We also performed a flow cytometry experiment where an EGFR positive cell line (U118) showed more than 1 log increase on the phycoerythrin spectrum scale when bound, compared to an unbound sample. We have been able to conclude that our aptamer is indeed selective, and preliminary experiments indicate that this selectivity remains when the aptamer is labeled with zirconium-89. In the future, we will attempt to reduce nonspecific binding in both fluorescent and radioactive binding assays by refining the aptamers synthesis and purification strategies. We will also begin testing the optimal aptamer constructs in our in vivo xenograft models. These xenograft experiments will include growing cancer cell lines in culture, injecting these cells into nude immunodeficient mice to grow the xenograft tumors, followed by tumor imaging with the Zr-labeled aptamer.

Mycobacterium tuberculosis (MTB) and nontuberculosis mycobacteria (NTM) are inconsistently identified in nonhuman primate (NHP) laboratories. Infections by MTB and NTM are common in NHP colonies and can interfere with immune-based diagnostic assays. Accurate and prompt detection, identification, and treatment are essential to understanding how tuberculosis (TB) research conducted on NHPs can be applied to human populations. The laboratories of Gerard Cangelosi and Lisa Jones-Engel recently validated Oral Swab PCR (OSP) as a cheaper and faster alternative to sputum-based diagnostic methods in human patients. OSP utilizes real-time PCR-based detection of the multi-copy IS6110 insertion sequence in MTB DNA, and therefore does not require an immune response. This project aims to 1) characterize the presence of MTB in rhesus macaques using polymerase chain reaction (PCR) based detection; and 2) to test the sensitivity of OSP in nonhuman primate laboratory populations that serve as biomedical models in many fields of research. OSP was performed on 320 primate swab samples from the Oregon National Primate Research Center (ONPRC) with and without histories of TB infection. These data were compared with OSP results to validate OSP as a means of TB detection in NHP. Following this analysis, multiple samples collected from confirmed disease-free (N=50), immunocompromised (N=50), and MTB-infected (N=20) animals at ONPRC will be evaluated. Furthermore, OSP will be evaluated as a means to detect NTM. If validated in this and other projects, OSP can not only provide a better understanding of disease exposure and bacterial transmission dynamics in NHP models, but also in human populations.

Epileptic encephalopathies (EE) are severe forms of epilepsies where recurrent seizures contribute to significant neurocognitive impairment and developmental delay for children with this disorder. Recent studies have shown that some cases of pediatric EE are caused by copy number variants (CNV), which are chromosomal deletions, duplications, or insertions of DNA across the genome. CNVs may contain specific genes that are important for neural health, some of which can cause disease when there is a copy number change. Despite significant advances in epilepsy gene discovery, the majority of pediatric EE cases remain unresolved. I am investigating whether I can identify rare CNVs in EE patients where the genetic cause is not known. I will screen EE patients for CNVs using a technique called array comparative genomic hybridization (aCGH). This technique will identify specific loci where a pathogenic, or disease-causing, CNV may exist in a patient that is not found in the control genome. I anticipate that if a CNV is found in an EE patient, I will determine its pathogenicity by comparing the genotype and phenotype of the patient to parents tested by aCGH to determine if the CNV is de novo - arising for the first time in that individual and not inherited from either parent. We also expect that in families who have experienced seizures or have seizure disorders, patients may inherit rare mutations from either parent. Finding a genetic basis for epilepsy by identifying rare mutations will advance our understanding of the pathology of the disease. Furthermore, identification of particular pathogenic genes will be useful in the clinical setting for genetic counseling of affected patients and families.

The amino acid tryptophan can participate in biological reactions as a reducing agent, in which a single electron is transferred producing a tryptophan radical . In these reactions, tryptophan radicals can participate in electron transfer chemistry, such as in DNA photolyase or catalytic chemistry, such as in the peroxidase enzymes. The properties of tryptophan radicals are heavily dependent on the microenvironment made up of the surrounding protein. The objective of our research is to study how the protein environment can control the properties of tryptophan radicals. This is accomplished by making controlled changes to a model protein called the beta hairpin, which contains tryptophan. In order to study the effects of these controlled changes, spectroelectrochemical techniques are being developed. In situ EPR (electron paramagnetic resonance) spectroelectrochemistry is a highly desirable technique for generating tryptophan radicals within an EPR spectrometer, giving simultaneous access to cyclic voltammograms and EPR spectra. The electrochemical technique cyclic voltammetry utilizes a three electrode system to measure the current carried in a sample while sweeping across a voltage range. In EPR, a frequency is a held constant while sweeping across a magnetic field range. The absorption is measured, and the resulting spectra provide information about the environment surrounding the tryptophan radical. Implementing cyclic voltammetry within the limited space (sample tube inner diameter of 1.0 mm) of an EPR sample tube presents multiple challenges. The process in designing such a cell has been developed and implemented in the study of tryptophan radicals.

 Kelp are a family of macrophytic algae that are important ecosystem engineers and key components of marine productivity. Light plays an important role in the lifecycle of kelp as an energy source and as an important growth regulator. In kelp haptera (the root-like structures that attach kelp to substrate) light induces a negative phototropic response. A negative phototropic growth response is where an organism grows in the opposite direction of a light source. In the ocean, shorter wavelengths of light penetrate water deeper than longer wavelengths. This produces an effect where the full spectrum of visible light can be available at shallow depths, but only blue light is available at deep depths (with gradient of wavelengths). Thus it was hypothesized that kelp species generally inhabiting shallower water (e.g., Alaria marginata) would have a phototropic response to red (720-740 nm), green (520-540 nm), and blue (420-440 nm) light. Species generally inhabiting deeper water (e.g., Agarum fimbriatum) would have a stronger negative phototropic response in the blue wavelength of light in comparison to other color treatments. Kelp haptera from both species were isolated and exposed to red, green, and blue light for 16 days. The controls were exposed to unidirectional sunlight or no light. The hypothesis was mostly confirmed. The blue light exposure caused the highest phototropic response in both species, but had a higher relative response in A. fimbriatum. Alaria marginata haptera responded to all wavelengths of light, refuting literature citing that haptera only respond to blue light. Green exposure produced the lowest response in both species. Moreover, it was discovered that red light was responsible for an increase in the number of haptera in A. marginata. The ecological implications and mechanism behind this red light driven haptera growth are opportunities for further research.

4,4-diaminodiphenylsulfone (DDS) is an antibiotic commonly prescribed for treatment of leprosy. Because leprosy was once believed to be highly contagious, patients in South Korea were quarantined on Sorok Island off the country's southern coast. It was later found that the patients treated with DDS had a longer average lifespan relative to the general Korean population. DDS has also been reported to delay aging and extend lifespan in the roundworm Caenorhabditis elegans, but the mechanism by which this is achieved is currently unknown. This project's purpose is to identify the pathways and mechanisms involved in lifespan extension from DDS treatment in C. elegans. Whole genome RNA sequencing and quantitative real-time PCR were performed to determine which genes were differentially expressed in DDS treated C. elegans. From the results of the RNA-sequencing and qRT-PCR analysis, we were able to observe that among the most highly induced genes under DDS treatment were genes that coded for the enzymes hsp12.6, a type of heatshock protein that is expressed in response to heat stress, hsp12.3, and fmo-2, a flavin containing monooxygenase. Through other experiments we have also observed that DDS fails to significantly extend lifespan in hif-1 (hypoxia inducible factor) knockout mutants, suggesting that hif-1 is important for lifespan extension from DDS. To further validate our RNA-sequencing and qRT-PCR data, we are currently using green-fluorescent protein reporters to visualize the expression of hsp 12.3, hsp 12.6, and fmo-2 in DDS treated C. elegans and performing experiments to measure the lifespan of C. elegans that overexpress fmo-2. Thus far we have identified several candidate genes involved in lifespan extension from DDS treatment and have formulated an incomplete but developing model that describes DDS' mechanism of action. 

In order to survive, cells must be able to regulate growth in response to changes in their environment. The Target of Rapamycin kinase Complex (TORC1) is one of the key regulators of cellular growth and division, and abnormal TORC1 signaling is found in multiple types of cancer and disease. TORC1 is turned off an on by Rheb, a small GTPase. The activation and regulation of Rheb within the mammalian TORC1 pathway is well understood, but little is known about how Rheb is activated in C. elegans. Given that nematodes lack identifiable homologues of the mammalian Rheb regulating proteins, understanding Rheb activation in this system could shed significant light on the mechanism and relevance of nematode growth in response to nutrients. To find genes that regulate Rheb we conducted a mutagenesis screen that revealed two different worm mutants (ENU9 and ENU10) with abnormally high Rheb expression. Using rapid single nucleotide polymorphism (SNP) we mapped the approximate genetic locations of the mutations in both strains. To confirm these results, we used the ENU10 mutation’s chromosomal location, which was obtained using fluorescent mapping strains. Analysis of whole-genome sequencing has suggested possible candidates in this genetic region. This work brings us closer to identifying key genes that affect Rheb activity in the nematode.

In 1994, the North American Free Trade Agreement (NAFTA) came into effect, slowly removing certain tariffs on exports and imports between Mexico and the U.S. Officials who supported free trade argued that NAFTA would significantly decrease Mexican immigration to the U.S by creating jobs in Mexico. This research project examines and measures the direct relationship between NAFTA and the increasing number of laboring immigrant communities developing in Washington’s apple producing counties. Personal accounts, articles, and documentaries used in our research indicate the necessity of Mexican apple growers and workers, to immigrate to the U.S in search of jobs. Census and archival records recognize a substantial Mexican population growth since 1994 throughout Washington State’s primary agriculture counties. Today, Washington State accounts for 85 percent of all apples imported by Mexico. As a result, Mexican apple producers are unable to compete in their own market with the subsided and highly efficient Washington apple industry. In effect, Mexican apple growers are forced out of business and paradoxically the market for Washington apples expands. In turn displaced Mexican apple growers and workers increasingly immigrate to Washington in order to answer the expanding market demands in the state. The interplay between NAFTA and Washington’s apple industry is fascinating because as exports to Mexico increase so does the demand for Mexican legal and undocumented workers. Areas like Yakima, Wenatchee, and Chelan have experienced up to 168 percent Latino increase from pre-NAFTA to early 2000s. Our research attempts to connect the strong development of laboring immigrant communities in apple producing counties as consequence of NAFTA policies that create economic and social problems for apple growers in Mexico. The final outcome of all this, is the rise of a sometimes vulnerable and dependent Latino communities working under harsh, and exploitive circumstances.

 In patients with multiple sclerosis (MS), both cognitive impairment and physical pain can have a severe effect on quality of life. This study explored the influence of pain on objective neuropsychological impairment in patients with multiple sclerosis, and compared that to the influence of pain on subjective cognitive complaints in MS patients. Data for this study was taken from neurocognitive exams administered to 127 patients with MS. Pain was evaluated on a 1-10 scale. To evaluate objective cognition, the following tests were administered to patients: California Verbal Learning Test (CVLT), Wechsler Adult Intelligence Scale (WAIS-III), Paced Auditory Serial Addition Test (PASAT), Symbol Digit Modalities Test (SDMT), Controlled Oral Word Association Test (COWAT), and Trail Making Test (TMT). Results showed that changes in pain intensity did not significantly predict changes in scores of any neurocognitive test administered (p>0.05) besides the COWAT (p=0.038). To assess whether the same trend was present between pain intensity and perceived cognitive ability, a different set of data was consulted. This data was taken from neurocognitive exams administered to 398 patients with MS. Conversely to objective cognitive scores, pain intensity scores significantly predicted changes in perceived cognition (p<0.05). These results suggest that, while improvements in pain through treatment may not improve objective cognitive impairment, they may help patients accurately predict their level of cognition.

Ionic liquids are chemicals that are both liquid at room temperature and contain atoms with formal charges. These two properties make ionic liquids excellent solvents; not only can they dissolve both polar and non-polar compounds, but their effect can also be easily turned by mixing them with water. This makes ionic liquid solutions very good at facilitating many nanoscale processes, namely protein-catalyzed ones. Since proteins normally can only perform their usual function in a water based environment similar to that found in living cells, the discovery of a new medium in which these proteins can function has a myriad of interesting nanofabrication applications. To study the viability of ionic liquid solutions as protein solvents, several computer simulations have been done on the short model peptides LK-alpha-14 and LK-beta-15. These peptides were exposed in silico to both pure water and ionic liquid solution while at an air-solution interface, and simulated for 50 ns. Preliminary results show that the peptides keep their secondary structure, which makes them highly likely to retain their function. This is promising, as it shows that ionic liquid solutions can successfully solvate these proteins without denaturing them, unlike many other less polar solvents that have been tried. With this information, future simulations can now be done that will hopefully show actual processes working with important protein catalysts, such as enzymes.

Childhood obesity has more than doubled in children and quadrupled in adolescents in the past 30 years. In 2012 it was estimated that 18% of children and 21% of adolescents were obese in the United States. Obese adolescents in particular are likely to be prediabetic and are at greater risk for poor physical and mental health. Moreover, obese youth are likely to become obese adults. In recent years, mindfulness-based interventions targeting eating behaviors such as binge eating, portion sizing, and emotional eating have emerged. It is hypothesized that by cultivating mindfulness—or the quality of moment-to-moment consciousness—health behaviors can be improved. This study aims to review mindfulness interventions in obese populations. A literature review was conducted to identify studies that describe the use of mindfulness in the context of obesity-related eating behaviors. Papers were collected from the databases Google Scholar, PubMed and PsycINFO using a combination of the following keywords: mindfulness, eating, obesity, adolescents, weight, intervention, school intervention, emotional eating, food, and awareness. Of the 39 papers reviewed, three frameworks were identified based on research methods: interventions (n=24); concept analyses (n=10); and articles teaching mindfulness (n=5). While the majority of papers addressed the role of mindfulness in obesity (n=38), only seven addressed youth specifically. Among these were three interventions for adolescents (one with obesity, another with Prader-Willi syndrome, and a feasibility study for preventing obesity in urban youth) and four articles for teaching mindfulness to youth. The interventions showed that mindfulness positively influenced weight control and eating behaviors including binge eating, emotional eating, and portion sizing. Mindfulness interventions, while not thoroughly described, have been shown to be effective, particularly for improvement of eating behaviors (i.e. stress eating, food cravings, and body awareness). More research on mindfulness interventions, specifically mindful eating, for obese youth is recommended.

There has been an increasing desire to reduce the number of research animals and shift to in vitro alternative methods to test developmental neurotoxicity as for many other types of toxicity testing. To study developmental neurotoxicity, the previously published in vitro micromass culture of rat embryonic midbrain cells was applied to mouse embryonic midbrain cells. The objective of the study was to characterize in vitro C57BL/6J fetal mouse midbrain micromass culture. The fetal mouse midbrain cells were isolated and cultured for 21 days after plating. Morphology was examined throughout time, and the cells were stained with hematoxylin for determining neuronal differentiation days in vitro (DIV) 1, 2, 4, 6, 8, 15, and 22. The result of the hematoxylin staining showed that the midbrain cells become more differentiated over time, but the intensity of staining may not be reliable once the culture goes beyond DIV 15 because the area of micromass expands to the sides of the plates which lead to the decreased intensity of staining. Mouse midbrain cells were also harvested for proteins at DIV 1, 2, 4, 6, 8, 15, and 22. Proteins increased over time, indicating not only differentiation, but also proliferation of the mouse midbrain cells. This was also confirmed by western blot and immunohistochemistry by characterizing the expression of neuronal-specific markers such as PCNA and nestin for proliferation, β-tubulin III and MAP-2 for differentiation, and midbrain specific marker, En1. Based on these results, proliferation may be at its highest at DIV 4-6 in culture system while differentiation is greatest in DIV 6-8. This mouse midbrain micromass culture system is a good model to investigate developmental neurotoxicity.

Urban watersheds have been shown to have excess nutrient loading due to a multitude of factors, including storm water runoff containing fertilizers and animal waste.  These nutrients often lead to eutrophication and the loss of recreational uses in urban lakes and waterways.  Wapato Lake in Tacoma, Washington, has had a history of toxic algae blooms as a result of excess nutrients.  An alum treatment was applied in 2008 in an attempt to limit the availability of phosphorus which is a key factor in algae blooms.  This study continues a monitoring program measuring the nutrient and hydrologic budgets of Wapato Lake.  The purpose is to assess the likelihood of harmful algae blooms and to provide information to guide future managment.  Water samples were taken from both the North and South basins over the course of a year and analyzed to calculate phosphorus and nitrogen reservoirs and fluxes.  Total numbers of waterfowl on the lake were determined at the same time as the water samples were taken.  Zooplankton and phytoplankton relative abundances were determined.  Water levels were measured over time by pressure gauges placed in the lake verified with physical readings on staff gauges and corrected for atmospheric pressures from a nearby station.  This data is used to construct a nutrient mass balance model to be used to aid managment programs that aim to improve future recreational use of Wapato Lake.

Gene delivery has successfully been accomplished with the use of polycations, both in vivo and in vitro. However, a major problem in this category is the inherent cytotoxicity associated with a high density of positive charges. Polymers that can be degraded within the cells are typically better tolerated compared to their non-degradable counterparts. This research studies the use of a dibromomaleimide linking agent to reversibly connect cationic polymer moieties to a stable polymer backbone. Reducible and non-reducible, statistical co-polymers of 2-aminoethyl methacrylate (AEMA) and oligo(ethylene glycol) monomethyl ether methacrylate (OEGMA) were synthesized and evaluated for a comb-like, backbone chain. These polymers have been chosen due to their low cytotoxicity as well as their high degree of polymerization control in reversible addition-fragmentation chain transfer (RAFT) polymerization. Backbones of DP 50 and 100 have been successfully synthesized. The current phase of the project is to functionalize these polymers with dibromomaleimide followed by reaction with 4-(dimethylamino)thiophenol to yield the reversible polycation material. This final polymer will be used to transfect cells with genetic material to test its feasibility for gene delivery. Success of this project will create a well-defined polymer containing positively charged cations that are easily dispersed upon internalization by cells.

This abstract is no longer available. Please contact urp@uw.edu if you have any questions.

Cancer cells usually develop resistances to therapeutic drugs due to high genetic and phenotypic heterogeneity. Our research focuses on targeting a more genetically stable tumor associated macrophage (TAM) populations in the tumor microenvironment. Macrophages are broadly classified as either M1 cells that enhance killing of pathogens or M2 cells that promote tissue repair. TAMs have M2-like properties which promote tumor progression. Our lab has identified a fusion peptide, M2pepKLA, which contains a macrophage-targeting M2pep sequence and an apoptosis-inducing KLA sequence. However, rapid degradation of this peptide in serum prevents it from exhibiting its full therapeutic potential in vivo. Our initial experiment reveals a specific site in the M2pep sequence that is readily cleaved by serum proteases. In this work, we aim to improve the serum stability of M2pep by rational substitution of the susceptible amino acid residues. The modified M2pep analogs will be evaluated for binding to M2 macrophages to ensure the binding activity is not compromised. Then, we will test the peptide stability against degradation in mouse serum. Our initial trial on D-amino acid substitutions failed to retain M2 macrophage-binding property. We are now investigating the effect of alanine and tyrosine substitutions as a means to improve the peptide stability. Lastly, the optimized construct will be fused with KLA to evaluate its efficacy to target and deplete M2 cells in complete media in vitro. We expect to derive a serum-resistant M2pepKLA analog that binds comparatively well to M2 macrophages and enables their depletion even in the presence of serum.

Neurotransmitters are chemical signals that mediate communication between neurons and other cells of the body. One type of neurotransmitters, modulatory neurotransmitters, such as dopamine, insulin, serotonin, and norepinephrine, are stored and released by dense core vesicles (DCVs). Due to the content of the vesicles, DCVs play important roles in managing human behaviors, emotions, and appetite. Therefore, dysfunctions in pathways controlled by neurotransmitters carried by DCV are commonly related to diseases, such as schizophrenia, depression, drug addiction, eating disorder and diabetes. So far, five proteins, RAB-2 and four of its effectors, have been identified to be essential for the formation and maturation of DCVs. In this project, I, in the Ailion lab, attempt to study the interactions between RAB-2 and its effectors by performing in vitro binding assay using purified proteins.RAB-2 is a GTPase proteins, which alternately bind to guanosine triphosphate (GTP) and guanosine diphosphate (GDP). When they bind to GTP, the proteins are active, able to recruit their effectors, and able to function. Meanwhile, in their GDP bound state, the proteins become inactive. Beforehand, to ensure that this method will work, I have been making controls for this method based on other well-known RAB interaction systems. In this experiment, I expressed and purified RAB-3 with its effector, UNC-10, and RAB-27 along with its effector, RBF-1. Thereafter, I will perform the binding assay by immobilizing GTP and GDP-bound RAB proteins to glutathione resin in separate tubes. Then, I will incubate them with their known effectors. I am hoping that, indeed, using this technique, the experimental result will give good positive controls, showing that RAB-3 will only bind to UNC-10 and not to RBF-1, and RAB-27 the other way around. Then, I will continue my project to examine the interactions of RAB-2 with its effectors using the same manner.

Our ever-changing external environment demands from us the ability to successfully switch between behaviors when cues indicate that it is appropriate to do so. The capability to respond to external, reward predictive cues is particularly important for species, as this is often key for survival. While these cued switches in behavior are routine for the majority of people, various psychopathologies including major depressive disorder, generalized anxiety disorder, drug addiction, and obsessive compulsive disorder result in an impairment in this skill, warranting research into the underlying neural mechanisms that allow an organism to perform this critical behavior. Recently, animal studies investigating the processing of reward have focused on the lateral habenula (LHb), a midbrain structure critically involved in learning and decision making. While the majority of prior research describes the LHb as an anti-reward signal in which its neurons fire upon the absence of an anticipated reward, some studies have provided evidence indicating that the LHb may be important for more than strictly negative reward prediction errors. Through muscimol inactivation and neuronal recording, the present experiment aims to understand how the LHb contributes to decision making when advantageous behavior is signaled first by auditory cues, and secondly by the presence or absence of a reward on a maze based task. Male Long-Evans rats learned that auditory cues (high- or low- pitched tones) signaled which of the two arms in a figure-eight maze contained a sugar pellet reward. Cues were switched every 3-6 trials of the program, requiring the rodents to repeatedly and proactively switch their behaviors in order to receive reward. Preliminary data collected upon inactivation of the LHb prior to task performance resulted in severe task impairment, suggesting that the LHb may act as a cue processor, important for encoding reward predicting cues in addition to reward prediction errors. 

Recent lab studies suggest that endophyte-assisted phytoremediation can be an alternative way of disposing and degrading organic pollutants. In this study, we selected a naturally occurring endophyte called PDN3 to degrade trichloroethylene (TCE), a suspected carcinogenic organic pollutant found at many industrial waste sites. The PDN3 endophyte will be inoculated in a hybrid poplar called OP367. The colonization of PDN3 in OP367 will be confirmed using fluorescent microscopy. A field study has already been conducted in a TCE contaminated site and the results clearly indicate that the poplar trees inoculated with PDN3 were much healthier than those that were not inoculated. This study is aimed toward testing if the PDN3 endophyte is responsible for the increased degradation of TCE by measuring the chloride concentration in soil of PDN3 inoculated and uninoculated poplar trees. According to our understanding, we should see a larger concentration of chloride concentration in the soil of the inoculated plants, thus inferring that more TCE is being degraded and promoting plant growth.

Since the discovery of the RNA interference (RNAi) pathway in 2001, short double-stranded sequences of RNA known as small interfering RNA (siRNA) have been studied extensively as potential therapeutic agents for cancer treatment. SiRNA causes post-transcriptional gene silencing via mRNA degradation, and can thus suppress overexpressed proteins that are essential for cancer cell survival and proliferation. However, siRNA-based therapies have not yet achieved clinical translation, in part due to poor cellular uptake and trapping within endosomes after uptake. We have designed a tetrameric fusion protein for siRNA delivery that addresses these issues and should improve overall gene silencing. The tetramer consists of four recombinant fusion proteins, each with three domains: a double stranded RNA (dsRNA)-binding domain, an endosomal escape peptide, and a tetramerization domain. This design provides siRNA stabilization and charge shielding, endosome disruption through the proton-sponge effect, multivalence for increased cellular uptake, small size for deep tumor penetration and non-immunogenicity. Cancer cell targeting is achieved through the enhanced permeability and retention (EPR) effect and cancer receptor-binding aptamer RNA sequences attached to siRNA. The tetramer is synthesized using molecular cloning and Escherichia coli expression systems, and its binding ability, size, and purity are characterized using blue native PAGE (BN-PAGE) and dynamic light scattering (DLS). The tetramer’s gene silencing capabilities are tested in vitro on LNCaP cancer cell lines using free and protein-bound GAPDH-silencing siRNA. Silencing is quantitatively assessed based on cellular mRNA levels using reverse transcription real-time PCR (RT-qPCR). The tetramer-bound siRNA and free siRNA are discussed in terms of their relative silencing and endosomal escape capabilities. If successful, this tetramer will be one of few protein-based delivery systems for siRNA, and could greatly improve the potency of siRNA cancer treatments.

Levonorgestrel intrauterine system (LNG IUS) is a highly effective contraceptive that has decreased the rate of unintended pregnancies in the United States. While overall rates of discontinuation are relatively low, most common reasons for method discontinuation are bleeding and pelvic pain. The purpose of this study is to develop and test a standardized anticipatory counseling clinical intervention using video that addresses benefits and expected side effects, such as bleeding and pelvic pain, associated with device use. We will measure the video’s effect on women’s rates of continuation and satisfaction with the LNG IUS. We predict that women exposed to the LNG IUS video compared to a control, unrelated video, will be better informed in regards to LNG IUS expected side effects and will have lower rates of discontinuation. The study will be conducted in two phases. The first phase of the study deals with video content development and production. Health care professionals and prospective LNG IUS users will be asked to provide feedback on the video in terms of accuracy, credibility, and clarity. The second phase of the study deals with the implementation of the video in the clinical setting and an evaluation of the effectiveness of the video in regards to method satisfaction and duration of use. It is anticipated that patients in the intervention group will report higher satisfaction and continued usage of the LNG IUS. Findings of this study can be used to determine the effectiveness of video to counsel patients and encourage adherence to treatments. Increasing evidence on the efficacy of video for counseling purposes can revolutionize physician-patient consultations and possibly substitute the widely used pamphlet or time-intensive in-person counseling.

"A Glass Half Empty: The Absence of Female Politicians in Bosnia-Herzegovina (BiH)" is the result of analyzing data gathered during a research trip to Bosnia. After an intensive course in Bosnia-Herzegovina's geopolitical history, myself and nine other students accompanied two professors to research transitional justice efforts in 2014. In ten days we interviewed eleven organizations, traveled to four different towns, and visted historical sites and genocide memorials.This research summarizes the terror and resolution of the Bosnian War (1992-1995), specifically focusing on its impact on women. Amongst other horrors, the war resulted in more than 100,000 people dead and more than 20,000 women raped. The U.S. brokered Dayton Peace Accords of 1995 did not bring peace for the survivors of sexual assault, nor any plan for the empowerment and envolvement of the women in the country's future. Many scholars believe that women are "paid lip service" in the constitution, but actually have little opportunity for political involvement. While conducting research, I became keenly aware of the strong representation of women in BiH's civil society sector. Many of the NGOs that we visited were led or influenced by women, all of whom were highly educated and committed. Women make up the majority of students in BiH's universities, and have a strong track record for implementing innovative programming for local peace-building organizations. However, all of these positives were profoundly at odds with the weak representation of women in national politics. The paper explores the contradiction that exists between the educated and active women enriching BiH's civil society, and the absence of such crucial voices in BiH politics. It also examines the ways in which women are absent from politics—including candidacy, elected officials, and significant legislation—and probes what factors have contributed to the silencing of women in the political sphere.

Contamination of the local environment is often a byproduct of industrialization. Although steps have been taken to prevent future contamination, there remains a need to address a legacy of industrial negligence. Phytoremediation offers a natural, affordable method of remediating these sites by taking advantage of the ability of certain plants to tolerate and accumulate environmentally harmful agents as biomass. Endophytes, non-pathogenic microbes that live within plants, maintain a mutualistic symbiosis with their host. We are investigating whether this relationship allows the host to endure a wider range of conditions that would otherwise be toxic; including habitats containing elevated concentrations of toxic heavy metals. We isolated individual endophytes from the microbiome of plants living in arsenic contaminated soil, then selected for arsenic tolerance from among these microbes. Arabidopsis thaliana seeds inoculated with a pure culture of Pseudomonas spp., designated PD9R, were grown on solid media supplemented with either 50 uM or 100 uM sodium arsenate (AsV). The inoculated plants showed higher germination rates, increased dry biomass and longer root length compared to the uninoculated controls. This increased resilience suggests promising applications in the use of endophytes for the phytoremediation of polluted soils.

Acetamiprid is a neonicotinoid pesticide used on vegetables and fruits to prevent insect damage. The mechanism of action for neonicotinoids is activation of an organism’s nicotinic acetylcholine receptors in the nervous system. Activation of these receptors can cause excitatory neurotoxicity, potentially leading to acute poisoning, paralysis, or death. While neonicotinoids have relatively low mammalian toxicity, some cases of acute acetamiprid toxicity have been reported in humans. Human exposure to acetamiprid can result from such activities as direct handling of pesticides, contact with treated surfaces, or eating produce containing chemical residues. While neonicotinoid exposure can be determined by indentifying chemical metabolites in human urine, research in this area has been limited, especially for metabolites specific to acetamiprid. In this study, identification and quantification of acetamiprid metabolites was investigated in human urine. Previous studies have looked at the recovery of individual acetamiprid metabolites. This study investigates the simultaneous recovery of the parent compound, acetamiprid, and two metabolites: acetamiprid-N-desmethyl (ACP-N-desmethyl) and 6-chloronicotinic acid (6-CNA). Urine samples were processed using Solid Phase Extraction (SPE) and the compounds quantified using high-performance liquid chromatography and tandem mass spectrometry (HPLC-MS/MS). A variety of SPE parameters were investigated in order to optimize metabolite recoveries including cartridge media and eluting solvent. The method was tested on urine samples collected from pesticide handlers after applying acetamiprid in an orchard to determine their occupational exposure. Initial method development results have shown acceptable recoveries of the acetamiprid parent compound and ACP-N-desmethyl from human urine. These metabolites show promise for use as biomarkers to identify acetamiprid exposure.

After a shock wave crosses an interface with plasma, the shock speed along with its shape change. These changes are dependent on the geometry of the interface and the plasma temperature. The goal of this project is to comparatively study how the shock dispersion on thermal inhomogeneity is affected by a plane and spherical geometries of the interface. The results of new findings can be used in for aerodynamic flow control in supersonic flights by creating plasma around a moving vehicle, in the detonation theory and combustion through the control of the ignition conditions, and for environmental improvements through sonic boom reduction.

Our lab is investigating the possibility of a magnetic dipole orientation sensor that is compatible with biological samples. Utilizing a thin near-surface layer of nitrogen-vacancy centers in diamond, we are able to optically image small magnetic field disturbances. This has allowed us to detect and locate magnetic nanoparticles on the diamond surface under ambient conditions. In this work, we examine the feasibility of using a similar sensor to detect the magnetic orientation of the particles, in addition to their location. I have developed software to simulate the imaging system, taking into account factors such as the size and material of the particles, as well as the optical properties of the sensing surface. I will present simulation results indicating that orientation measurements should be possible, as well as initial experimental results for 50 nm and 500 nm magnetic particles that were deposited on a diamond surface. The ability to detect the orientation of biologically compatible magnetic nanoparticles will likely be useful in the study of a wide variety of microbiological processes, enabling the tracking of complex motion on the nano-scale.

Most bees are solitary and differ from social bees (e.g. honeybees) in that every female bee is fertile and independently maintains her own nest. All bees are important pollinators and depend entirely on flowers for food: pollen provides proteins, lipids and vitamins, while nectar provides energy. To determine when and how much pollen adult female alfalfa leafcutting bees consume, bees collected during the summer flight season were dissected to evaluate the relative quantity of pollen within each of the three major regions of the digestive tract. Bees were found to consume pollen throughout the day and flight season. Furthermore, accumulation of pollen in the crop (before passing into the midgut for digestion) showed patterns intermediate to those of other studied solitary bee species; this suggests that pollen passage through the crop might be associated with bee nesting biology.

MicroRNAs are a type of noncoding RNA transcript involved in the regulation of gene expression in eukaryotes. Recently, a number of microRNAs have been heavily implicated in colorectal cancer, including miR-21, miR-145, and miR-20a, among others. These microRNAs have been found to be present and dysregulated not only in the solid tissue mass of tumors, but also circulating in the blood plasma of colorectal cancer patients. Here I outline the initial steps for creating a non-invasive diagnostic and prognostic test for colorectal cancer screening through the use of highly sensitive RT-qPCR (reverse transcription quantitative polymerase chain reaction). Previously, we have designed highly sensitive and specific RT-qPCR primers and assays for the successful detection of subclinical levels of HIV in blood serum. We now turn these same primer design techniques to cancer detection. The particular assay being implemented involves the use of custom-designed stem-loop RT primers for the reverse transcription and cDNA synthesis of microRNA target genes of interest within a multiplex reaction utilizing highly sensitive TaqMan DNA probes. By screening for low amounts of circulating microRNAs, we hope to not only predict the presence of cancerous cell growth, but also to be able to track cancer progression, response to chemotherapy, and even defining of cancer sub-types.

Women make up only 25% of employees in the technology industry (e.g., computer science careers, information technology, etc.). One explanation for this disparity is that hiring decisions are made based off cultural fit, or the perception of similarities between employers and job candidates. In previous research, we discovered no difference in hiring intention between male and female candidates, but male candidates were seen to fit better into the company environment than female candidates. The current research investigates whether perceptions of cultural fit are determined in part by the stereotypicality of the candidate, and if males are seen to fit better in computer science (CS) environments even when females express the same interests and appearance. In our study, undergraduates were asked to imagine that they were hiring a job candidate for a technology company. Participants read a description of either a non-stereotypical (e.g., art posters on the wall, etc.) or stereotypical (e.g., Star Trek posters, computer parts, science fiction posters, etc.) CS workplace, and then read a stereotypical (e.g., hobbies include Star Trek Trivia, Anime club, etc.) or a non-stereotypical (e.g., hobbies include intramural soccer, Music appreciation club, etc.) resume of either a male or female job candidate accompanied by a picture of the candidate. Participants then answered questions measuring perceived candidate fit into the company environment and hiring intention. We predict that stereotypical candidates will be seen to fit better into the company environment and be hired more often than either of the non-stereotypical candidates. We also predict that while stereotypical female candidates will fit better than non-stereotypical female candidates, they will fit less well and be hired less often than stereotypical male candidates. This would suggest that perceptions of the overall stereotypicality of the candidate prevent non-typical candidates from being hired, and may limit women from entering the industry.

This presentation is focused on the limited research conducted to date on non-electronic gaming as an alternative speech and social therapy method. It also explains the benefits shown in the results of available data and suggests research to be conducted in the future. The past decade has seen shifts in the academic community to the usefulness and applications of video games. That view is not held, however, for the non-electronic gaming counterparts. There has been very limited research conducted on the practical uses of these types of games, especially in the field of psychology. None of these studies directly sought to use these as tools for therapy, despite results suggesting that non-electronic gaming offers improvements not seen in electronic games. The much stronger social dynamic of face-to-face intentional interaction, as well as the necessity of more complex speech, are categorically different than electronic games. Results have shown improvement in: motivation, facilitation of effective group work, enhanced trust, willingness to reciprocate, and speech impact. All of these desirable skills can be difficult to treat at times in traditional therapy models. Some research is beginning to take place in educational fields, with the most applicable studies to date being centered on those with Autism Spectrum Disorder. While useful, there is need to expand the research further, to seek implications with other populations. Additional research must be conducted to not only develop useful therapies, but to better understand the mechanisms underlying their effectiveness. This is a topic uniquely suited to psychology in comparison to other fields. Research need not be limited to removing all electronic components though, as potent results are seen in electronically aided face-to-face pervasive gaming. The electronic components of these formats all have much stronger social interactions that are not seen their traditional video games counterparts.

The goal of our research is to enhance online health communities for caregivers and patients managing cancer by making it easier to reach out to mentors for advice and support. Exploration and outreach in online communities can be hindered by the physical and emotional stresses of the cancer experience. Moreover, current online resources often leave users feeling overwhelmed. Accordingly, users would benefit from new tools or frameworks for gathering information and connecting with others in online environments. Using social matching theory, we developed a way to match users with potential mentors based on samples of written text. We are now evaluating these recommendations through a user study with patients and caregivers to learn what factors people value in a good mentor and what information helps users choose peer mentors. To begin, we ask participants to write a short story about their cancer experience. The story is used to generate a set of potential mentors from online community data that we represent in paper profiles. We then conduct a follow-up session where participants rank mentor profiles and answer qualitative questions about their perception of online communities. Several recurring sentiments have already been observed. Cancer patients and caregivers alike placed importance on “give and take” mentor/mentee relationships and time efficient interactions. Demographics, diagnosis, and sample posts have also been favored when assessing the potential of a mentoring relationship. Moving forward, we will run a statistical analysis on profile rankings and affinity diagram qualitative data. The insights acquired through this study will inform new designs that align the functionality of online health communities with user needs and values.

We are interested in filling in the puzzle of how cooler night temperature affects the genes involved in the flowering time in Arabidopsis thaliana. CONSTANS (CO) and FLOWERING LOCUS T (FT) are important genes which integrate several environmental cues to regulate flowering time. CO protein is a key transcriptional factor activating FT expression, which in turn induces flowering. The ambient environment light cycle coincides with the temperature cycle, resulting in an induction of CO mRNA and FT mRNA at the dawn. However, the interactions among the genes controlling flowering on the molecular level are still blurry. We made the prediction that cooler night temperature suppresses both CO and FT expression. In the experiment, we treated Arabidopsis with three different conditions: LD 22/22, LD 22/12 and LD 12/12. LD stands for long day with 16 hours of light and 8 hours of darkness. We harvested the seedlings every four hours starting from dawn for RNA isolation, cDNA synthesis and used qPCR to calculate the amount of CO and FT expression. To eliminate the differences between individual seedlings, we normalized with constitutively expressing gene, IPP2. Analyzing the qPCR data, we found that seedlings under LD 22/12 condition had an obvious increase of CO expression during the night. Compared to FT level in the seedlings under LD22/22(control), FT level under LD 22/12 treatment is suppressed at dusk and higher at dawn. As for LD 12/12, CO does not show strong induction during the night; however, FT stays at a high expression level throughout the day. The different patterns in the alterations in CO and FT expression profiles suggest that even though CO is activating FT expression, other environmental cues, such as the diurnal temperature cycles coinciding with the day length cues, are regulating FT level. In the future, we are interested in the upstream regulatory processes that drive these responses.

The study of horizontal stripe development in zebrafish is used to understand the cellular and molecular basis of pattern formation. The zebrafish adult stripe pattern consists of three pigment cell types: melanophores, iridophores, and xanthophores. Previous work identified that the zebrafish seurat mutant arises from mutations in the gene encoding Immunoglobulin superfamily member 11 (Igsf11). seurat (igsf11) mutants develop spotted patterns rather than stripes due to defects in melanophore migration. Three separate alleles have been identified in igsf11 showing similar phenotypes however none are predicted null alleles. The CRISPR/Cas9 (Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR associated nuclease 9) system is an effective means of gene modification and knockout. Here we use this system to target igsf11 in wildtype zerbafish with the goal of generating a complete loss of function allele. Through this technique we aim to further characterize the role of Igsf11 in melanophore migration during adult stripe formation. Zebrafish embryos were injected with an igsf11 gRNA and Cas9 RNA. We verified that these injections induced deletions in igsfll by using HRM (high resolution melt analysis). Additionally, we observed defects in melanophore stripes in our injected fish, suggesting biallelic igsf11 mutations. Through classic complementation testing to screen for germline carriers we crossed injected fish to seurat (igsf11) mutants and looked for offspring with spotted patterns, suggesting non-complementation with igsf11. This method of screening was highly successful and most of the potential carriers we screened had germline mutations in igsfll. To identify the causative mutations, we sequenced igsf11 near the gRNA target site. Overall, we found that CRISPR/Cas9 is an effective system for generating mutations in specific genes of interest. Now armed with additional alleles of igsf11, we can test for interactions with other genes involved in zebrafish melanophore migration and patterning.

Mitochondria are essential organelles that are most commonly known for their pivotal role in cellular respiration via oxidative phosphorylation. Additionally, mitochondria are involved in other important cellular processes such as lipid biosynthesis and apoptotic cell death. Mitochondria have dynamic properties including fusion, division, and movement, which are all essential to maintain mitochondrial activities. Given the plethora of essential cellular pathways that require mitochondria and their products, it is not surprising that defects in mitochondrial function are implicated in many different human diseases and disorders including myopathies, diabetes, and neuropathies. In mammals, Mitofusins 1 and 2 (MFN1 and MFN2) mediate outer-mitochondrial membrane fusion. Dysfunction in MFN2 has been linked to some Charcot-Marie-Tooth neuropathic disorders including Charcot-Marie-Tooth type 2A (CMT2A). Mutations associated with CMT2A are found in each of the various functional domains encoded by the Mfn2 gene. In addition to its role in outer-mitochondrial membrane fusion, MFN2 may have a role in such diverse cellular functions as mitochondrial-ER interplay, working with the MAVS protein in innate immunity signaling, and in mitochondrial transport. Here we will generate representative mutant versions in each functional domain of MFN2 and characterize their rescue activity in mammalian mfn2 knockout cell lines. In characterizing the activity of these mutant proteins, we hope to gain insight into the pathophysiology of CMT2A and the mechanism of MFN2 function.

An estimated 2.3 million people live with multiple sclerosis (MS), an immune-mediated disease, worldwide. People living with MS can experience a wide range of potentially debilitating symptoms that affect how they engage in society. The Financial Assistance Program offered by the National MS Society is intended to provide guidance, support, and resources to help manage the financial impact of MS. The goals of the intervention are to encourage independence, safety, health, and quality of life for clients. The aim of this study is to evaluate the impact of the Financial Assistance Program offered by the National MS Society, Greater Northwest Chapter, on clients living with MS and their families and friends. A survey including scaled and open-ended questions assessing the perceived impact of the program was utilized to gather data. Outcome measures included self-efficacy in advocating for needs, awareness of community resources, independent living, and perceptions of impact on family and friends. Quantitative and qualitative analyses offer a nuanced understanding of the impact of the Financial Assistance Program. Clients indicated a notable difference in perceived impact between critical short-term needs and more sustainable forms assistance. Financial assistance eased burdens on clients’ friends and families. The National MS Society, Greater Northwest Chapter, has many support groups, and there were requests for additional social support groups made by clients. Qualitative evidence suggested that support group members act as ambassadors within their communities to spread awareness about MS and the mission of the National MS Society. Furthermore, additional engagement of clients’ families and friends could broaden the impact of the Financial Assistance Program. This program evaluation will inform improvement and expansion of the Financial Assistance Program at the National MS Society, Greater Northwest Chapter, as well as identify areas in which advocacy from the organization is needed at the policy level.

Our laboratory recently showed that interhemispheric callosal connections correlate closely with ocular dominance columns (ODC’s) in the primary visual cortex (V1) of pigmented Long Evans rats. In the lateral segment (LS) of V1, callosal connections form a homogeneous band, whereas in the binocular, central segment (CS), callosal connections form distinct patches that correlate only with ipsilateral ODC’s. Moreover, the LS in Long Evans rats is monocular, responding mainly to visual stimulation through the contralateral eye. In contrast, preliminary electrophysiological data suggest that the LS in V1 of albino rats is binocular rather than monocular. This surprising difference between rat strains led us to hypothesize that albino rats lack segregated ocular dominance domains in V1. We hypothesized that: (1) ipsilateral and contralateral eye-specific projections are intermixed in V1 of albino rats; (2) callosal connections in V1 of albino rats are not patchy as in Long Evans rats, but rather are distributed homogeneously and project a mix of signals from both contralateral and ipsilateral eyes; and (3) the binocularity found within the LS of albino rats depends directly on input from the ipsilateral eye relayed by callosal projections from the contralateral CS. Our current electrophysiological and anatomical findings are consistent with these hypotheses. In addition, we sectioned the corpus callosum to test the hypothesis that callosal connections provide input from the ipsilateral eye to the LS in albino rats. Neurons in the LS of callosotomized albino rats mainly responded to stimulation through the contralateral eye, providing additional support for our hypotheses. Our study points to the albino rat as a useful model for studying factors that regulate segregation of eye specific input in the mammalian visual cortex.

In this project I will research different micro-RNA sequences found in serum pertinent to the diagnosis and prognosis of Hepatocellular Carcinoma, which is the most common type of liver cancer. Micro-RNA sequences are used in the post-transcriptional regulation of mRNA and gene expression, and have been particularly studied for their roles in cancerous tissues/serums. I will attempt to create TaqMan assays using real-time PCR to observe increased or decreased expression of certain micro RNA sequences in HCC afflicted serum samples. Ultimately, my project aims to determine if selected micro RNA sequences are significantly up-regulated or down-regulated in HCC positive serum samples to reflect the presence of HCC as a biomarker and to determine its use as a prognostic tool.  

When crises, including natural hazards, school shootings, and political unrest, occur people attempt to make sense of the events occurring around them. This sense-making process often results in ‘rumors’ (information of unknown validity). With the growing adoption and use of social media these rumors travel faster and reach farther than ever before.The massive amount of information circulating at any given moment makes it is difficult to distinguish rumor from fact, and it is often impractical for a human (e.g. emergency responders, citizens, etc.) to systematically differentiate the two. In response, our research aims to design an algorithm which can identify potential rumors without human supervision. We hypothesize that rumors have unique patterns in both their linguistic and temporal structure, which (when correctly identified) will allow for automated detection. Our current work uses data from Twitter collected during multiple crisis events; data also include multiple rumors (stories) which have been identified by human coders. By combining quantitative methods from the fields of social network analysis, computational linguistics, and machine learning we design and test rumor detection algorithms (using the human-identified rumors as a benchmark). Our current algorithms have succeeded in identifying many of the rumors in our test data, but a high rate of false-positives remains a prime issue. Moving forward we would like to further explore temporal patterns of rumors, meta-data patterns, and noise reduction techniques for our current methods.

Plants have evolved to use day-length information to coordinate flowering time with appropriate condition to maximize their reproduction by regulating expression FLOWERING LOCUS T (FT). In Arabidopsis, blue light receptor FLAVIN-BINDING, KELCH REPEAT, F-BOX 1 (FKF1) forms a complex with GIGANTEA (GI) under presence of light to stabilize CONSTANTS(CO), an activator of FT expression. The FKF1-GI module conserved from early lineage of land plants, such as non-vascular bryophyte, determines the timing of reproductive transition by sensing the length of daytime. It suggests that FKF1-GI is one of the highly conserved systems to induce long day specific responses of plants. There is growing evidence showing that longer day-length is critical to induce senescence and oxidative stress responses both can be regulated by cellular redox status. Those responses are enhanced by increasing light intensity especially under long day condition. Also, gi mutants showed longevity phenotype under oxidative stress condition, suggesting that GI, potentially together with FKF1, might be involved in regulation of oxidative cell death processes. Likewise, FKF1-GI can modulate various long day specific responses, and the role of FKF1-GI could be more important under long day coinciding with higher light intensity. Therefore, it is worthy to test the function of FKF1-GI in regulation of long-day specific responses. In order to achieve such goals, firstly, we grew wild type (WT) Arabidopsis plants under different light intensity to test the effect of light intensity on known FKF1-GI function (i.e. flowering time regulation) and other long day specific response (senescence), and set the experimental condition for further approach. Under physiological range of light intensity (80~500 umolm-2s-1), flowering of WT plants were slightly accelerated by growing light intensity. We also checked the degree of senescence of WT under different light intensity. By analyzing anthocyanin and chlorophyll contents, we found that WT plants under higher than 300 umolem-2s-1 of light intensity had an obvious early senescence phenotype. In case of fkf1 and gi-2 mutants, even under higher light intensity (500 umolm-2s-1), there is no difference compare with that in low light intensity. It suggests that FKF1-GI could be a master switch to induce photoperiodic flowering and senescence responses of plants, potentially for completing reproduction of plants within given seasonal time tables.

The Maly Lab works primarily with protein kinases in order to characterize their functions and produce highly selective and potent small molecule inhibitors. My role in this research has been to synthesize small molecule inhibitors that can selectively and effectively inhibit Calcium Dependent Protein Kinase 1 (CDPK1). CDPK 1 has been shown to be essential for the invasion of Toxoplasma and Cryptosporidium into mammalian cells. Toxoplasma and Cryptosporidium are human pathogens for which new therapies with increased efficacy and reduced toxicity are needed. I am generating inhibitors based on a 4-amino pyrazolopyrimidine scaffold, and attaching different functional groups that can interact with unique regions of CDPK1’s ATP-binding site. This 4-amino pyrazolopyrimidine scaffold is structurally similar to adenosine triphosphate (ATP) without the phosphate groups. Therefore, these compounds are able to inhibit CDPK1 by competing with the ATP substrate. Functional groups were added by Suzuki couplings, acylations, and alkylations. None of my synthesized compounds have undergone biological testing yet, but similar molecules have shown inhibition at concentrations as low as 1 nM. Once the first set of compounds are tested, the most effective inhibitors will have addition functional groups attached to try to create more potent and selective inhibitors. It is important for these inhibitors to be very specific because there are over 500 different protein kinases in the human body and causing inhibition of the wrong kinase can have lethal effects. By the end of this project, our CDPK inhibitors will be selective and potent enough for medical use in combating Toxoplasma and Cryptosporidium.

The DNA repair protein O⁶-methylguanine-DNA methyltransferase (MGMT) is a known mechanism of resistance to alkylating agent-based treatments for human malignant gliomas, including current radiotherapy with temozolomide, the contemporary standard of care. The usual clinical methods for determining tumor MGMT content (e.g., immunohistochemistry) are limited by insensitivity and the association of surrogate measures of MGMT gene expression (e.g., CpG methylation status of MGMT promoter) with MGMT activity is not quantitative. The biochemical assay of MGMT has the advantages of being conceptually simple and relatively easy to perform. The assay exploits the fact that MGMT acts by transferring a methyl group from the O⁶ position of guanine and covalently binding it to a cysteine in the proteins active site. This mechanism produces native guanine in DNA and an inactive methyl modified protein.  By using exogenous DNA containing ³H-labelled O⁶-methyguanine, it is possible to estimate the number of MGMT molecules in a volume of tumor homogenate.  While biochemical assays circumvents the problems associated with other methods of determining MGMT content, it is seldom employed, even in laboratory settings, primarily because of the absence of a standard, optimized assay. The goal of this project was to establish a standardized assay that is optimal for determining activity in homogenates of glioma tissue.  To meet this objective we have: (I) Established basic biochemical parameters by determining A. linearity with amount of extract assayed and incubation time B. lower limits of detection C. reproducibility between assays D. effect of repeated freeze-thawing on extract activity. (II) Examined possible confounding factors found in tissue homogenates by determining the effect on activity of A. iron from blood on the re-dox state of cysteine B. salt concentration C. tumor DNA. Suppression of the glioma DNA repair capacity has the potential to lead to the development of new therapies that can dramatically improve the prognosis for individuals with malignant gliomas.

Bacterial pathogens activate a complex set of signaling pathways that facilitate adaption to distinct environments within the host during infection. Salmonella typhimurium possess RpoS: an alternative sigma factor for RNA polymerase. RpoS directs gene expression programs that help Salmonella adapt to environmental stresses including starvation, high osmolarity, and oxidative stress. As such, RpoS regulates multiple genes that may play a role in the regulation of the flagellar gene expression cascade. This cascade is composed of three classes; class I and II encode transcriptional activators required for the expression of the subsequent class. I investigated the role of RpoS in the regulation of flagellar class I, II, and III genes in Salmonella typhimurium laboratory strain 14028. I hypothesized that RpoS has multiple regulatory effects on the flagellar gene cascade: RpoS will regulate the transcription, translation, and stability of class I genes which in turn will affect the expression of class II and III genes. To test this hypothesis, I constructed three strains: 14028 ++RpoS, 14028 +RpoS, and 14028 ΔrpoS using the lambda red system. I performed single-cell analysis using GFP reporter constructs and flow cytometry, multi-cell analysis using β-galactosidase assays, and western blot analysis to determine the level of expression of class I (flhDC), class II (fliA), and class III (fliC) flagellar genes under varying levels of RpoS in both stationary and exponential phase. Based on this data, I can conclude that RpoS has both positive and negative regulatory roles at different stages in the flagellar gene cascade.

The optical spintronics and sensing lab researches the fundamental properties of defects in semiconductors for quantum-enabled technologies. The lab is extending its measurement capabilities to the wide bandgap material, ZnO, which requires a near ultraviolet laser source. Here we will use second harmonic generation (SHG), a common type of nonlinear optical interaction, to realize a laser at 369 nm from incident laser light at 738 nm, a wavelength readily available in the laboratory. The SHG laser source consists of a birefringent crystal chosen from BBO (Beta Barium Borate), BIBO (Bismuth Borate), and LBO (Lithium Triborate), all of which are common non-linear optical materials that are often used for SHG. Design parameters include the crystal’s nonlinear susceptibility, its thickness, and cut angle. The cut angle is determined by phase matching between the fundamental wavelength and second harmonic to optimize the SHG efficiency. Birefringence exhibited in these crystals allows us to achieve this phase matching because we are able to find a propagation direction through which both wavelengths experience a same refractive index. The crystal design and efforts toward experimentally realizing the SHG source will be presented.

There is an urgent need to develop new therapeutic drug nanocarriers to treat infections caused by airborne exposure to biothreats, specifically Francisella tularensis and Burkholderia pseudomallei. These two biological weapons are highly infectious, aerosolizable, intracellular alveolar pathogens and have been identified by the Centers for Disease Control and Prevention (CDC) as CDC Tier 1 select agents. The intracellular compartmentalization of these pathogenic organisms within alveolar macrophages is a significant barrier to bacterial clearance and contributes to their associated morbidity and mortality. Currently, there is no licensed vaccine available and very few antibiotics such as ceftazidime, ciprofloxacin, and doxycycline have proven to be effective in treating these pathogens only under prolonged antibiotic therapy.This project will develop a novel polymeric nanoparticle that can deliver multidrug combinations intracellularly to targeted alveolar macrophages and overcome pathogens’ drug resistance mechanisms. Initial studies have demonstrated the feasibility of preparing nanoparticles with controllable hydrodynamic diameters with molar mass dispersity values as low as 1.10. Based on these initial experimental successes the project is now focused on optimizing conjugation chemistries in order to synthesize nanoparticles containing with precisely defined concentrations of two antibacterial agents. These drugs, which are linked to the nanoparticle via ester bonds, can be liberated from the nanoparticle scaffold via a combination of hydrolysis and enzymolysis in acidic endosomal compartments. The next phase of this project will be to evaluate the rate of drug release from the nanoparticles under physiologically relevant conditions. This project will also evaluate different drug combinations and nanoparticle formulation parameters in order to establish the optimum antibacterial composition. In vitro evaluation of the efficacy and toxicity of the nanoparticles in intracellular bacterial clearance in alveolar macrophages will be conducted through co-culture, MTS, and planktonic assays.

Macrophages of our immune system can be broadly classified as pro-inflammatory M1 macrophages or anti-inflammatory M2 macrophages. Chemical factors lead to a localized imbalance in macrophage phenotypes that have been associated with various chronic diseases, suggesting that targeting specific populations of macrophages may achieve therapeutic benefit. Phage display is one method of identifying peptides that can be used for targeting cell populations. In whole-cell peptide phage display, a library of bacteriophages, in which each phage displays a unique peptide, is applied to target cells. Phage that preferentially bind target cells are isolated, and their peptide sequences can then be obtained by DNA sequencing. Targeting human M1 macrophages has possible clinical applications in detecting and treating arthritis, diabetes, and atherosclerosis. In this work, I used phage display to identify two novel phage-bound peptide sequences, named phM1.1pep and phM1.2 pep, that exhibited selective binding to M1 macrophages. ELISA and flow cytometry binding studies suggested that these two phage-displayed peptides exhibited higher binding to M1 over M2 macrophages and overall higher binding than insertless phage that lacked displayed peptides. My next step was to extract a binding peptide sequence from the phage by synthesizing various peptide designs and evaluating their binding to M1 and M2 macrophages. I initially synthesized only the displayed peptide sequences and compared binding selectivity to M1 macrophages between peptide designs having C-terminal or N-terminal tags (for visualization). In previous work, it was shown that the peptide tag location had a major influence on peptide binding. I also assessed whether adding amino acids adjacent to the displayed peptide on the phage coat enhanced binding to M1 macrophages. In future work, I aim to explore applications of these peptides by conjugating therapeutics to the peptides for targeted delivery to M1 macrophages.

The majority of cognitive tests used in mouse research are modified versions of those originally designed for rats. This tailoring has led to recent concern as to whether such cognitive tests are truly optimized for mice. The purpose of this study is to evaluate the ability of a Radial Water Tread (RWT) maze to differentiate between Traumatic Brain Injury (TBI) and Sham subjects in mouse models. TBI represents a significant health risk among veterans, and optimization of accurate cognition and behavioral tests for mouse models of TBI is essential to the development of efficacious treatments. 10-week-old male C57BL/6J mice (n =17) were randomly assigned to receive a Sham (8) or Controlled Cortical Impact (CCI) surgery (9). Mice were tested using the Grid Test and Noldus Catwalk Gait Analysis at one-day post-injury and again at 32-days post-injury, with the addition of the Rotarod test. Starting on 12-days post-injury, mice were tested using the RWT maze. Each subject was given 3 trials a day, for four days (acquisition period), with short-term and long-term memory test on the fifth (STM) and twefth day (LTM), respectively. Trials lasted a maximum of 180 seconds, and scores were averaged across trials. Results revealed the RWT maze was successful in distinguishing between Sham and TBI at both STM (Shams=20.81±14.68 & CCI=48.58±37.66, p<0.05) and LTM (Shams=10.86±8.10 & CCI=70.54±57.79, p<0.05) time points. Catwalk showed significant differences at both TBI post-injury time points over multiple variables. Grid Test analyses are ongoing. Rotarod analysis had no statistical significance. These data validate the use of the RWT maze as a credible cognitive test for mice in TBI models. Future research will examine a closed-skulled CCI (CS-CCI) model, which provides a more realistic model of human TBI and could provide a greater understanding of the consequences of head injuries.

Ectopic calcification (EC) is the process of abnormal mineralization of soft tissues and current approaches for treating EC have limited efficacy. Cell therapy utilizing osteoclast cells—bone remodeling cells in our body—could provide an alternative treatment for EC. In Giachelli Lab, a cell therapy that allows differentiation of osteoclasts from genetically engineered monocytic precursors (RAW264.7+iRANK) in a controllable manner has been developed. However, a method for delivering the cells is still needed. In this study, a hydrogel vehicle was proposed to assist delivery of the osteoclast precursor cells. Three biodegradable hydrogels were proposed for this study: type-I collagen, fibrin, and Matrigel. Cell-hydrogel interaction and cell viability were investigated through in vitro 2D and 3D experiments, where cells were seeded underneath and encapsulated within the hydrogel. Via Von Kossa staining, we demonstrated that osteoclast precursor cells could resorb mineral on 2D surfaces when seeded under each of the three gel conditions. Using live/dead staining and Tartrate-resistant acid phosphatase (TRAP) activity assay, we also showed that cells could still proliferate and differentiate when encapsulated in gel. In addition, we demonstrated that our in vitro TRAP activity results corresponded to in vivo study in nude mice (data not shown), where collagen encapsulated cells and no gel control performed equally well, and fibrin encapsulated cells showed maximum amount of cell differentiation. Future study would include post-delivery analysis of hydrogel encapsulated osteoclast precursor cells in differentiating and bone resorbing ability. The findings of our in vitro model would assist the delivery of an osteoclast cell therapy developed by Giachelli Lab and advance the field of engineered cell therapy.

Social media are changing the way people socialize with others, communicate with friends and family, identify and express themselves, and collaborate. As social media continue to become integrated into everyday life they become core platforms for self-presentation and identity formation, as well as social influence. Indeed, many social scientists now recognize the valuable research opportunity that such environments present for unobtrusive observation of human behavior. In this project, we use social media data as an indicator of alcohol and drinking-related behaviors in young adults. While many have claimed that digital traces is predictive of offline behavior, we are able to directly explore these claims using a valuable dataset of self-report survey responses of behavior from 215 participants in combination with observations of behavior, more than 300K posts, on the micro blogging platform Twitter. We address the basic question of do young adults discuss their drinking behavior on social media? By utilizing text analysis methods, we aim to gain a better understanding of whether social media can be an indicator of young adults' risky alcohol-related behavior. This research has important implications for identifying at-risk young adults. It also has the potential to inform the design and implementation of health-related intervention strategies for youth.