Humans are incapable of regenerating a majority of their major organs and tissues following traumatic injury, often resulting in an irreversible loss of function. Tadpoles of the frog genus Xenopus can regenerate multiple tissue types in response to injury, however this capability is lost after metamorphosis. This stage-specific regenerative capacity makes Xenopus a uniquely powerful model for studying factors that promote regeneration. Tadpoles develop ex-utero and do not develop mouths until days after fertilization. Before tadpoles are able to ingest exogenous food, they rely instead on maternal yolk stores for sustenance. A regenerative refractory period has been described in tadpoles of Xenopus laevis, in which regenerative capacity is transiently lost. In this study we describe a similar refractory period in the closely related Xenopus tropicalis, and observe that the onset of the refractory period aligns with the transition independent feeding. Based on this observation, we hypothesized that the lapse in regenerative capacity could be due to a lack of metabolic fuel. We used immunohistochemistry (IHC) against the yolk protein vitellogenin (vit) to study the utilization of maternal yolk stores during tadpole development. We find that yolk localization is dynamic over the course of development, and that it is ultimately is depleted by the onset of the refractory period. We additionally used IHC against phospho-Histone 3 (pH3), a marker of mitosis, to study proliferation during development and regeneration. We found that proliferation declines across development heading into the refractory period, in both uninjured and amputated contexts. Lastly, we successfully rescued both regeneration and proliferative rates by feeding tadpoles after they develop the ability to eat. As a whole, this work articulates that nutritive stress may contribute to the loss of regenerative capability in the refractory period, and that alleviation of this stress promotes regenerative ability in this context.

The direction that plants grow in is dictated by the directions of gravity and light. A mutation in the LAZY-2 (LZ-2) gene in tomato (Solanum lycoperiscum) causes the plant to actively grow downwards, in the direction of gravity. It has been shown that the lz-2 phenotype is dependent on the phytochrome (Phy) protein family; a group of plant proteins that is responsive to red light. In a plant with no functional phytochromes, the lz-2 mutation is restored to wild-type, and the plant will grow upwards as normal. In this project, we discern the roles of three of the five tomato phytochromes, A, B1, and B2, to see if they work together or alone to provide the light cue that initiates the mutant function of lz-2. By gravistimulating combinations of lz-2 and Phy tomato seedlings under red light, we have detetermined that PhyA delays the lz-2 response, while PhyB2 exhibits an additive effect with PhyA and PhyB1 to effectively reverse the lz-2 phenotype. These data expand on previous work to further our knowledge on the relationship between light and gravity sensing in higher plants. 

Wildfires are known for their destructive capacity towards ecosystems and devastating impacts to human life and property. Regions in the western United States are particularly prone to such events, including large crown fires. However, there is limited research on the potential for smoke from these wildfires to carry and redistribute nutrients in the form of aerosols. Here I show that higher levels of airborne nutrients, specifically iron, phosphorus, and potassium, can be associated both with the timing of active wildfires and a known smoke tracer, elemental carbon. Data from the Interagency Monitoring of Protected Visual Environments (IMPROVE) show a strong correlation with fire presence and local peaks in levels of potassium, with less strong associations with phosphorus and iron. Across a multi-year period, there is also a correlation between the number of acres burned in a particular year and the average concentration of iron and potassium in that region. Using data from the NASA DC-8 airborne laboratory corroborates this information, with calculated potassium concentrations matching those on the ground. These findings indicate the wildfire plumes have a potential to be significant sources of nutrients in the short term. This is especially relevant for areas impacted by wildfires, as these nutrients are key for plant growth and development. These elements may also be carried into the ocean and affect the aquatic biosphere. All of the information gathered will help improve our understanding of the complex networks that make up Earth’s biogeochemical cycles.

Humans are incapable of regenerating a majority of their major tissues following traumatic injury. Tadpoles from the frog species Xenopus tropicalis have the ability to regenerate lost spinal cord, vasculature, muscle, and cartilage within a few days following injury. The regulatory mechanisms of gene expression necessary for regeneration have not yet been well defined. My primary interest is in understanding the relationship between stress signaling and gene expression during regeneration. The lab has shown that the stress responsive transcription factor Hypoxia Inducible Factor 1α (Hif1α) is necessary for the expression of Wnt target genes, one of the primary signaling processes necessary for regeneration. While we have found that Hif1α is necessary for Wnt target gene expression, we do not know the epistatic relationship between Hif1α and Wnt. In order to test this relationship, I utilized the drug IWR to antagonize Wnt and found that tadpoles treated with IWR have reduced tail regeneration 72 hours post amputation (hpa). I then supplemented these tadpoles with DMOG to stabilize Hif1α and found that DMOG is sufficient to rescue tail regeneration, suggesting that Hif1α is downstream of Wnt. In order to determine if Hif1α is sufficient for Wnt target gene expression, I extracted RNA from regenerating tails 24 hPa and used quantitative PCR (qPCR) to determine relative gene expression. I also utilized in situ hybridization to see if expression of these genes is restricted to regenerating tissues. As Wnt is a known regulator of neural and muscle development, I investigated how inhibiting Hif1α would impact complex tissue regeneration and found that Hif1α is necessary for regeneration of axons and muscle specifically. By determining the epistatic relationship between Hif1α and Wnt through the analysis of specific gene expression, we continue to improve our understanding of how regenerative organisms convert stress signals to cell fate signals.

A wide range of diseases and disabilities are associated with mitochondrial dysfunction including diabetes, cancer, Alzheimer’s disease, cardiovascular disease, and Parkinson’s disease. Mitochondrial dysfunction triggers activation of a novel mitochondrial retrograde response pathway in Caenorhabditis elegans termed the PMK-3 Retrograde Response, which upon activation leads to a reduction in mitochondrial stress and extension of lifespan. A mitogen-activated protein kinase (MAPK) cascade comprised of DLK-1, SEK-3, and PMK-3 forms the signaling core of the PMK-3 Retrograde Response. In this presentation, I will outline my attempts to develop photo-regulatable PMK-3 and DLK-1 kinases which are tools that can aid in the identification of other proteins involved in this pathway. These photo-regulatable kinases were made by engineering a recombinant form of the fluorescent protein Dronpa into two discrete sites of DLK-1 and PMK-3 to confer photosensitivity of the kinases. Photoregulation of the kinase occurs through dimerization of Dronpa in violet light (inactive kinase) and dissociation in cyan light (active kinase). After determining the identities of phosphorylated proteins purified from nematodes exposed to cyan and violet light using mass spectroscopy, a comparative analysis between the two datasets will suggest which proteins are involved in the PMK-3 Retrograde Response. Further investigation into these proteins could elucidate the role of these proteins in mitigating mitochondrial stress.

Treatment of neurological disease has made little progress due to the inability of many therapeutics to access the brain environment. However, delivery vehicles like nanoparticles can allow therapeutics to overcome brain-specific biological barriers including the blood-brain barrier (BBB), the dense extracellular space (ECS), and cellular targeting. The ability of nanoparticles to overcome these barriers is influenced by surface properties which can be modified through the formulation process. One understudied parameter is the choice of surfactant, molecules which stabilize nanoparticle formation and likely form an interface between the nanoparticle and brain environment. First, we investigated the potential toxicity of several commonly used surfactants on brain cells and slices. We added surfactant solutions to mouse microglial cells (BV2) or cultured brain slices and assessed cell viability two days later with colorimetric assays. Our results showed that while surfactants cholic acid (CHA) and polysorbate 80 (P80) caused toxicity at high doses, they were nontoxic at the low doses involved with nanoparticle formulation. Other surfactants, including Pluronic® F127 (F127) and poly(vinyl alcohol) (PVA), were nontoxic throughout the tested dose range. Interestingly, although the F127 compound is nontoxic on its own, nanoparticles formulated with F127 reduced cell viability. This result was not observed with any other nanoparticle-surfactant combination. Confocal microscopy indicated higher intracellular accumulation of the nanoparticles formulated with F127 compared to all other formulations, suggesting that toxicity is mediated by nanoparticle internalization and surfactant choice. Finally, we used a live cell imaging technique to capture videos of the nanoparticle internalization process. Building off these results, ongoing experiments will evaluate several nanoparticle-surfactant formulations on their ability to accumulate within brain tissue after in vivo administration. Findings from this work will guide nanoparticle design for future clinical translation.

Hidden bias, also known as implicit or unconscious bias, affects attitudes, thinking, and behaviors in everyday interactions. It contributes to poor continuity and quality of care, and mistrusting relationships between health care providers and patients. Patients may not be treated equitably due to different identities (race, ethnicity, gender, etc) or different diseases (obesity, diabetes, hypertension, etc). These negative outcomes lead to health disparities and inequities. Despite this evidence, training strategies to detect and address hidden bias in patient-provider interactions are not well characterized and do not fully utilize innovative informatics and technology approaches. Can we leverage innovative technology to identify implicit bias from nonverbal cues in interpersonal interactions? Can we then provide feedback that raises awareness of those biases? The UnBIASED project will develop computational sensing tools to assess nonverbal communication signals associated with implicit bias and provide feedback to patients and providers. This approach could shape the next generation of training strategies for hidden healthcare bias. Documenting the range and utility of strategies in prior work upon which this innovative approach expands is important. To characterize existing training strategies for hidden bias and ways that technology can help, I report on a literature review of existing interventions and recommendations to combat implicit bias in clinical settings. Using dimensions, such as format of intervention- paper, technology, interactional (e.g., standardized patients), I characterize training strategies and their utility from prior work. Through this literature review, I aim to identify the gaps in existing work that illustrate opportunities for informatics and technology innovations for addressing implicit bias in healthcare. This review will provide practical insights for academic medical systems and programs on ways that technology can extend medical education curriculum to address implicit healthcare bias.