Autism spectrum disorder (ASD) is a neurodevelopmental disorder marked by repetitive behavioral patterns and challenges with social interaction. Gastrointestinal symptoms are a common comorbidity of ASD, and individuals with the disorder tend to have a distinct gut microbial community composition and circulating metabolomes. Elevated levels of gut-derived metabolite 4-ethylphenolsulfate (4-EPS) are associated with ASD mouse models and children with ASD. Administration of 4-EPS to conventional mice induced atypical myelination and anxiety-like behaviors. 4-Ethylphenol (4-EP), its precursor, is produced by gut microbiota before host-mediated sulfation; however, its microbial biosynthetic pathway remains unknown. We propose a pathway involving stepwise conversion of plant-derived complex polysaccharides to 4-EP. My project aims to identify a gut microbial enzyme that completes the first step of this proposed pathway: a hydroxycinnamoyl esterase. After extensive literature review and biochemical study, candidate enzymes from resident gut microbes were identified and selected using bioinformatic tools. In vitro and in vivo experiments will be used to assess their activity towards model and dietary substrates. Remaining substrate and product concentrations will reveal species and strain specific enzymatic activity and substrate uptake. If the results are negative, this bioinformatics to experimental analysis pipeline will be repeated on new candidate enzymes. This work complements ongoing lab investigations to demonstrate the complete enzymatic pathway in a single species. Elucidating the microbial biosynthetic pathway of 4-EPS will contribute to detangling the gut’s role in ASD, regardless of if it is causal to the disorder or simply a biomarker. More importantly, studying the biochemistry and metabolism of gut microbiota supports the efforts to fill fundamental gaps in understanding the gut-brain axis. 

All mammals experience a slowdown of cardiac pacemaker rate with aging. The main mechanisms to explain that phenomenon are related to alterations in the ionic currents that underlie the diastolic depolarization phase of the action potential. We have previously reported that pacemaker cells from old mice have reduced L-type calcium currents. We further explore the mechanism underlying that reduction, testing cell hypertrophy and alteration in the scaffolding of L-type calcium channels as potential mechanisms. To test for cell hypertrophy, we combined immunostaining and high-resolution imaging to map the HCN4-positive pacemaker region of isolated upper heart explants from young and old mice. We compared cell length, width, and area between young and old cells. We also determined these morphological parameters in HCN4-positive enzymatically dissociated pacemaker cells. We found no significant difference in cell dimensions or area between ages, ruling out hypertrophy as a potential mechanism. We used mass spectrometry to identify expression changes in scaffolding proteins essential for calcium channel organization at the plasma membrane. Through this approach, we identified a large reduction of caveolin 3 as a possible mechanism. Caveolin is a protein essential to forming signaling microdomains between calcium channels and other proteins. Using western blotting, we confirmed a 50% reduction of caveolin 3 in isolated pacemaker tissues from old animals. Using proximity ligation assay and super-resolution microscopy, we showed altered recruitment of L-type calcium channels into caveolae. Our findings suggest that the age-associated decrease of L-type calcium current is caused by a reduced insertion of these channels in caveolae.

The heart's primary function is to pump blood to supply oxygen and nutrients to the body. The biomechanical principles of the heart are determined by specializations at the organ, tissue, cellular, and molecular levels. Little is known about how these specializations have adapted to sustain high heart rates in animals with extreme biology, as is the case of the hummingbird, whose heart rate above 1000 bpm makes it the endotherm with the highest heart rate observed in nature. We hypothesize that the hummingbird heart has evolved several adaptations at all the abovementioned levels to i) generate fast firing rates, ii) optimize electrical-contraction coupling, and iii) sustain fast contraction-relaxation cycles. Using different histological and imaging approaches, we have started to characterize the architecture of the hummingbird’s heart for the first time in a research lab. To describe the overall dimensions and structure of the hummingbird heart, we generated CT scans and 3D reconstructions of iodine-labeled Calypte anna hummingbird hearts. To characterize the organization of the tissue, we present data using hematoxylin-eosin and lectin stainings in fixed paraffin-embedded slices of the hummingbird heart. Our preliminary results showed that hummingbird ventricles have a cell density of 110 cells per 5000 µm2, around 7-fold larger than mouse ventricles. Ventricular cells in the hummingbird are 8-fold smaller with a cross-sectional area of 41 ± 4 µm2. Hummingbird hearts also have a higher capillary density with 18.0 ± 0.6 capillaries per 2500 µm2. Our results provide a foundation for structural and functional characterization of the hummingbird heart at an organ, tissue, and cellular level while opening avenues for further investigation of extreme cardiac physiology.

Each year, approximately 84,100 adolescents and young adults (AYAs) in the United States are diagnosed with cancer. Beyond the physical challenges that come with cancer, they also face significant psychosocial barriers, including gaps in health literacy, limited access to professional mental health support, and uncertainty about the future. Addressing these unmet needs requires innovative approaches, and social media, particularly Instagram offers a unique opportunity to bridge the gap between traditional healthcare services and the psychosocial needs of AYAs. With 2 billion users, Instagram is widely used for connection and information sharing, making it a valuable platform for health-related support and education. This study qualitatively analyzes Instagram posts under popular pediatric cancer hashtags to: (1) explore how AYAS and caregivers use social media for health information and support, (2) identify barriers they face in healthcare, and (3) examine how social media can enhance health literacy and education. A direct content analysis of 300 posts was conducted using a newly created Instagram account to minimize algorithmic bias. AI tools, including ChatGPT, Perplexity, and Microsoft Copilot, helped identify commonly used hashtags, which were then cross-referenced on Instagram. The most frequently used hashtags include #childhoodcancer, #childhoodcancerawareness, #pediatriccancer, #stupidcancer, #fightlikeakid, and #morethan4. An iterative coding process, using sets of 5–10 posts, was employed to develop and refine a codebook based on existing literature. Posts were categorized by metrics, user profiles, content types, health-related quality of life, and social support. My coding team consisting of myself, and 3 other investigators will code the data using REDCap, with descriptive statistics analyzed in R Studio. Findings from this study will highlight how Instagram can serve as a powerful tool to improve health literacy, education, and mental health support for AYAs with cancer, ultimately bridging critical gaps in healthcare accessibility and education.

Cortical tracking, a method that examines how neural activity encodes the dynamic features of the incoming speech stimuli, allows for the study of naturally produced continuous speech. Successful encoding of acoustic features is fundamental for language processing and comprehension. Studies show that cortical tracking of at least some acoustic speech features is already robust in the first year of life. However, it is unclear whether bilingual infants exhibit enhanced cortical tracking of non-native languages compared to monolingual infants, consistent with the idea of having a "bilingual advantage" as suggested in prior research. To investigate this, we recorded neural responses from 11-month-old English learning monolinguals, English-Mandarin learning bilinguals, and two mature comparison groups of English monolingual and English-Mandarin bilingual adults, while they listen to naturally produced, continuous, infant directed speech using electroencephalography (EEG) in three conditions: English, Mandarin, and Vietnamese. Stimuli were presented at an overall level of 70 dB SPL in a sound-attenuated booth. Using a combination of machine learning and linear modeling (i.e., Multivariate Temporal Response Function approach), we analyze the EEG signals using a multivariate encoding model consisting of acoustic features including envelope, envelope derivative, word onset, and phoneme onset. We hypothesize that both bilingual adults and infants will exhibit enhanced encoding of acoustic features in Vietnamese compared to monolingual adults and infants, indicating bilingual advantage in processing a third language. Additionally, we anticipate the bilingual advantage to be more prominent in infants than adults. These findings will contribute to the understanding of how bilingualism influences neural encoding across different languages and provide neural evidence of bilingual advantage in processing and acquiring a third language. I participated in study design, recruitment, data acquisition and analysis.

Organs maintain consistent shape, form, and volume through complex processes, one of which is cell-cell adhesion. E-Cadherin, a key cell-cell junction protein, is critical for cell shape, arrangement, and tissue structure. In this study, I investigate the role of E-Cadherin in the morphogenesis of the Drosophila Malpighian tubules, a model system where I can manipulate E-Cadherin expression and use fluorescence microscopy to observe the effects on organ growth. Previous work involved fixing and staining embryos to track E-Cadherin localization using fluorescent imaging to measure its intensity. I will further analyze E-Cadherin localization spatiotemporally by constructing a fluorescent fly line for live imaging during development. I expect E-Cadherin concentration to increase during elongation and to be enriched in looped regions of the tubules. To assess the requirement of E-Cadherin in organ formation, I will reduce its expression using RNAi and degradFP, expecting significant developmental defects due to the protein's vital role in morphogenesis. These defects will be quantified by comparing changes in cell and organ shape in control and E-Cadherin-reduced tubules. Additionally, I will help develop Python tools for 3D image analysis, including cell segmentation, creating a 3D model of E-Cadherin in tubular cells, and extracting protein intensity. Developing these tools not only enables our work in these tubular organs but also allows for comprehensive image analysis of other tubular 3D organ forms. Elucidating the precise mechanisms behind cell behavior, shape, and cell-cell interaction has important human health implications and will enable work in many other fields such as cancer, regenerative treatments, tissue growth, and organ synthesis. 

While some species cannot persist in urban areas, coyotes (Canis latrans) thrive in cities in part thanks to their varied diet and creative scavenging. Urban coyotes consume more anthropogenic foods and have more diverse diets than wild populations, and the quality of the anthropogenic foods they consume varies with the landscape of the city. In addition, consumption of particular anthropogenic foods can bring coyotes into conflict with resident human populations. What are coyotes eating in Seattle, and what does their diet composition say about the specific urban environments they inhabit? DNA metabarcoding, a technique used to genetically identify the species present in a sample, provided an initial idea of coyote diet composition. However, the metabarcoding data lacks resolution for plants, invertebrates, and some anthropogenic foods. This study investigates the diet composition of Seattle’s coyotes through traditional scat analysis, building on previous metabarcoding work to identify key diet items. Traditional analysis allows for better identification of plant and invertebrate species via the identification of hard-items such as bones, exoskeletons and seeds, and can provide additional resolution where metabarcoding primers lack specificity. In particular, traditional analysis contextualizes the dietary role of chicken— the presence or absence of physical items such as feathers clarifies if coyotes are eating domestic chickens or anthropogenic foods. I estimate the percent composition of each item in a given sample and compare these results to the metabarcoding data in order to compare the strengths of traditional and genetic techniques for diet analysis. My anticipated results provide valuable information regarding the dietary role of invertebrates, the plants coyotes consume and disperse, and if coyotes are consuming domestic chickens— highlighting the advantages of traditional analysis used in conjunction with metabarcoding. These results will help refine the methods of omnivore diet research and inform action to prevent human-wildlife conflict.

Assessing how organisms respond to shifting climatic conditions is crucial in the era of climate change to predict species' resilience to environmental changes. This study aimed to explore the effects of heatwaves on grasshopper development and fitness. Specifically, I investigated the reproductive potential of two grasshopper species within the framework of a common garden heatwave experiment. In Spring 2023, we reared the grasshoppers under three heatwave intensity treatments, exposing each treatment group to three heatwaves during set developmental stages. Afterward, I dissected the preserved females frozen for analysis, quantifying the number of primary and secondary oocytes in their ovaries. Oocytes develop into eggs and as such are a metric of reproductive potential. I hypothesized that increased heat stress would result in a decline in fecundity. However, we did not find a significant effect of the heatwave treatment on oocyte count, suggesting any fecundity effects of heatwaves are via a different mechanism. Understanding how organisms respond to changing environmental conditions is key to understanding how ecosystems will change in the coming years, and is important for informing conservation efforts.

Multiepoch scintillation studies of pulsars shed light on the structure of the interstellar medium (ISM) by finding scattering screens that affect pulsar radio signals. PSR J0332+5434 has previously shown multiple scintillation arcs, indicating multiple scattering screens. My research analyzed new observations of PSR J0332+5434 to improve our understanding of its scintillation properties and determine the number and locations of its scattering screens along the line of sight (LOS). I analyzed over 30 high-cadence observations using the Green Bank Observatory’s 20m telescope using scintillation, secondary spectra with Scintools, and time-series Jupyter notebooks to generate dynamic spectra, secondary spectra, and time-series. My analysis revealed two scintillation arcs, indicating at least two scattering screens. When I combined these arcs with transverse velocity measurements, I detected a third scattering screen. Comparing my results to previous studies showed that two of the screens had been previously identified, but the third screen had not been identified. This means that PSR J0332+5434 may have at least five scattering screens: four previously identified and one new screen from this study. Furthermore, one of the arcs I observed is spread out and shows significant asymmetry. Only one arm is usually visible at a time, which shifts from left to right throughout my observations. This asymmetry could be caused by the variation in electron density in a region of the ISM along the LOS, causing the radio waves to refract. I plan to conduct more accurate observations using the Green Bank Telescope to investigate the refractive wedge causing this asymmetry and to identify any new scattering screens. Finding new scattering screens in the ISM—the interstellar gas clouds causing radio wave scintillation—allows us to develop better electron density models to improve pulsar distance measurements and improve our understanding of the Milky Way Galaxy’s structure.

Learner experiences are under-examined in environmental learning research. Our research consists of studies of experiential aspects of environmental learning by undergraduate researchers, conducted over three years, culminating in a focus on how community-engaged learning (CEL) fosters connections between social justice, ecological consciousness, and student well-being. Research questions we came to consider were: What connections are students drawing between social justice and ecological consciousness? How does engaging in community-based environmental learning affect students’ well-being? Methods such as coding, memoing, reflecting through learning diaries, whole-part-whole analysis, and group collaboration all contributed to establishing an adaptable infrastructure of undergraduate research (UGR) in experiential aspects of the course. Our findings on students’ connections between social justice and ecological consciousness revealed their thoughts about becoming advocates, or “leaning toward justice”, though they had diverse prior knowledge and experiences. Findings on the CEL experience within the large course with regard to well-being showed how students integrate environmental education with community engagement, particularly in addressing issues such as food insecurity, environmental justice, and language barriers for immigrant communities. Some key themes found were that CEL promoted personal growth through unexpected learning, connection to nature & emotional relief, and a sense of belonging in research participants’ experiences. The significance of this research has been to establish a way for undergraduate researchers to drive experiential learning research, and to find research outcomes about how learning experiences foster awareness of social and ecological justice, encouraging students to see themselves as advocates for change. 

Lys-R type transcriptional regulators (LTTRs) are one of the largest families of bacterial transcriptional regulator proteins with over 850,000 known members.  Many of these LTTRs are enriched in our gut microbiota, whose metabolic processes affect human health outcomes. LTTRs regulate gene expression through the binding of specific ligands to their ligand binding domain. Currently, less than 500 of them have been studied which represents a severe knowledge gap that conventional methods of characterization are unable to keep up with.  We aim to create a high throughput methodology to characterize LTTRs by their corresponding ligands that regulate gene expression. We are currently developing an assay to use chimeric LTTRs, or engineered LTTRs that share the same DNA binding domain yet a variable ligand binding domain. The use of chimeric LTTRs, which will all bind to the same DNA promoter, will potentially allow dozens of LTTRs to be tested in one assay. Our work thus far has demonstrated that chimeric LTTRs can be expressed in E.coli cells and purified using affinity chromatography and magnetic bead purification. We have also demonstrated that their ligand binding domains are functional and specific via differential scanning fluorimetry, and that their DNA binding domains are functional using an electromobility shift assay using SYBR green and SYPRO ruby dyes. Future work will explore their ability to regulate gene expression when their proper ligands are introduced with a substrate-induced gene expression reporter assay. Then uncharacterized LTTR candidates to be made into chimeras will be selected via a bioinformatic sequence similarity network analysis for assay piloting. If successful, this assay has potential to elucidate new metabolic pathways of our gut microbiota allowing for better understanding of their complex relationship with the human body.

Less than 10% of drugs successfully transition from preclinical to clinical trials, principally due to the inability of currently used 2-dimensional models to simulate the 3-dimensional structure and function of human tissues. To develop 3D in vitro models of human vasculature for more efficacious screening of anti-atherosclerosis drugs, I created a device for constructing a perfusable tissue containing a lumen by leveraging open microfluidic patterning methods developed by our group: suspended tissue open microfluidic patterning (STOMP). The device can be used to pattern tissue with a hollow luminal structure lined with endothelial cells, which can be perfused via hollow posts the tissue is suspended between. Using surface tension-driven flow, a liquid hydrogel precursor solution flows through the open microfluidic channel and around the two hollow posts. After gelling, the tissue anchors to the post, contracts away from the sides of the microfluidic channel, and the STOMP device is removed. By adding a second STOMP device that can surround the first tissue another cell-laden hydrogel can be patterned around the first tissue, encapsulating it. To form a lumen in cardiac tissue, I will pattern the inner region with human umbilical vein endothelial cells (HUVECs) in an enzymatically degradable polyethylene glycol hydrogel, surrounded by human induced pluripotent stem cell-derived cardiomyocytes in fibrin hydrogel. Enzymatic degradation of the core region will form a cavity through which HUVECs will remodel the cavity walls, forming an endothelial lining. I will assess lining formation by adding fluorescent dextran to cell media being perfused through the device and measuring fluorescence through confocal microscopy in the surrounding region over time, allowing me to evaluate the permeability of the membrane to compare with physiological values. This model can then be used to screen treatments for atherosclerosis to study how drugs interact with cells in a 3D microenvironment. 

Syphilis, caused by Treponema pallidum (T. pallidum), remains a significant global health concern, with increasing cases worldwide. Doxycycline post-exposure prophylaxis (Doxy-PEP) has emerged as a potential strategy to prevent infection. However, widespread use raises concerns about the possibility that doxycycline-resistant T. pallidum strains might emerge and spread. This issue is alarming since doxycycline is a second-line therapeutic for syphilis and is often used in patients with allergies to beta-lactams or when beta-lactams are unavailable due to shortages. If genetic resistance to doxycycline were to develop in T. pallidum, it could undermine the effectiveness of Doxy-PEP and further narrow the range of treatment options for syphilis. To address this concern, I developed a restriction fragment length polymorphism (RFLP) assay to detect potential doxycycline resistance mutations in T. pallidum. This assay analyzes the 16S rRNA gene region of T. pallidum where most likely mutations could develop based on the analysis of other resistant pathogens. The assay was optimized using three synthetic 16S rRNA gene constructs containing the resistance-associated mutations and DNA from a wild-type T. pallidum strain (Nichols) as controls. The presence of mutations in the amplified control DNA was assessed by restriction digestion with the AluI, RsaI, and SfaNI enzymes, which can selectively cut wild type and mutant sequences and reveal specific mutations. The analysis of 60 archived samples from syphilis patients collected in the US, Madagascar, Argentina, and Sri Lanka is ongoing. Results will provide data on the frequency of doxycycline resistance mutations in T. pallidum, if any are found in this selected group of specimens. Developing a rapid, cost-effective surveillance tool is essential for monitoring potential resistance and preventing treatment failures when doxycycline is used.

Infants and adults process fundamental sound attributes such as pitch, timbre, and loudness differently, but the underlying neural mechanisms that drive these differences remain unclear. Pitch is the perceptual attribute of sound that can be arranged from low to high as in a musical scale, while timbre differentiates sounds of the same pitch and loudness, such as different musical instruments. Timbre, related to the spectral distribution of frequencies, is perceived as brighter when high-frequency energy increases. In natural sounds, pitch and timbre often covary; for example, a flute’s sound is both high in pitch and bright in timbre. Our prior research showed that infants outperform adults without musical training in pitch discrimination in the presence of random brightness variations. One possible interpretation is that adults have learned the statistical covariation between pitch and brightness, leading to efficient coding but poorer performance when these expectations are violated. To investigate further, we recorded mismatch negativity (MMN)—a brain response to unexpected sound changes—using electroencephalography (EEG) in both 7-month-olds and adults. We measured responses to pitch changes in two conditions: 1) with random brightness variations and 2) without brightness variations. The results are consistent with our prior behavioral findings: infant MMN amplitudes were comparable in both conditions, whereas adult MMNs were larger without brightness variation. These results are consistent with our past behavioral findings and the interpretation that infants have not learned the statistical covariation between pitch and brightness, thus experiencing less interference when discriminating pitch in the presence of random brightness variation. This research offers insight into the differences between how the infant and adult brains process fundamental attributes of sound that are important for speech and music perception. My role in this project involved the acquisition of EEG data, data management, and dissemination of research findings.

Estrogen plays a key role in metabolic regulation including fat distribution and glucose homeostasis. Less understood are the differences of estrogen metabolism in different fat depots, and the conversion mechanisms that underlie the effects we see between two prominent isoforms of estrogen, Estradiol (E2) and Estrone (E1). E2 dominates during reproductive years, shifting to an increase in E1 post-menopause. These hormonal changes contribute to a switch in fat storage from subcutaneous to visceral depots, elevating the risk of metabolic diseases. My research investigates how differences in estrogen metabolism, mediated by cytochrome P450 aromatase (CYP19A1) and 17β-hydroxysteroid dehydrogenase (HSD17B1), influence glucose uptake in inguinal (IWAT) and epididymal (EWAT) white adipose tissue. Using ex vivo explants from C57BL6/J male and female mice, I treated IWAT and EWAT with E1, E2, an agonist of CYP19A1, and an HSD17B1 inhibitor. Glucose uptake was measured at baseline, after 24 hours, following insulin stimulation. Additionally, I performed RT-qPCR to quantify depot-specific expression of CYP19A1, HSD17B1, and related metabolic genes. Preliminary results show that IWAT exhibits higher baseline expression of CYP19A1 than EWAT correlating with IWAT also demonstrating greater insulin-stimulated glucose consumption. E1 treatment decreased glucose uptake in both depots, while E2 had minimal effect in IWAT. Most interestingly, E1 and the forskolin + HSD17B1 inhibitor combination significantly decreased glucose consumption. This suggests depot-specific metabolic adaptations driven by differences in estrogen metabolism. The differences between IWAT and EWAT in estrogen-mediated glucose regulation offers new opportunities to better understand the metabolic impact of E1 and E2 in estrogen metabolism and glucose uptake. Understanding these mechanisms could inform strategies for targeting adipose tissue to mitigate impacts of insulin resistance and obesity, especially for postmenopausal women. My contributions include conducting tissue treatments, measuring glucose uptake, and analyzing gene expression data.