Student Discovery Session Abstracts

Our goal is to provide more accurate non-invasive MRI methods for diagnosing cardiac pathologies. The methods proposed can replace the more costly and morbid invasive procedures currently used in the clinic. Current gold-standard technique in MRI uses a contrast agent injection which can only detect the end results of a myocardial infarction (MI) or a ‘heart attack’. Recently, we have developed techniques that can detect edema, which is a sign of tissue injury, in an earlier stage toward heart diseases. Unlike current techniques used in clinic, our techniques do not use any contrast agent, and are faster and more robust to motion. One breath-hold acquisition can generate images with significantly less artifacts with higher spatial resolution:16 time-varying images and a parametric map, which can indicate the regions of inflammation or injury. Thus, our technique can be used for the consideration of early interventional treatment for the patients.

Sex Differences in the Development of Familial Hypertrophic Cardiomyopathy-An Energetics Perspective

Camille Birch, University of Arizona

The R403Q mutation located in the Î±- myosin heavy chain is responsible for familial hypertrophic cardiomyopathy (FHC). It is a primary disease of the sarcomere and is the most commonly identified cause of sudden cardiac death in young populations. FHC male mice possessing the R403Q mutation develop left ventricular dilation and eventual cardiac dysfunction. However, female R403Q mice only show left ventricular hypertrophy without dilation or dysfunction. Adenosine monophosphate-activated kinase (AMPK) is a known regulator of cellular energetics and subsequently AMPK is activated in the hypertrophic state. This leads to a shift in metabolic processes which affects the efficiency of adenosine triphosphate (ATP) hydrolysis. It is predicted that female R403Q mice will have an increased efficiency of ATP hydrolysis compared to males in order to compensate for the energetically unfavorable conditions experienced by R403Q hearts.

NHERF-1 Functions as a Molecular Switch between Cell Migration and Proliferation in Glioblastoma

Ashley Chavez, The Translational Genomics Research Institute

Glioblastoma multiforme (GBM) is the most lethal of the advanced glial tumors. While chemotherapeutic agents have been developed to treat it, they have been ineffective in targeting the invading cells. This may be attributed to the overexpression and amplification of EGFR in glioblastoma. (>50 percent of the cases). In previous work,Â it has been established that the Na+/H+ exchanger regulatory factor 1 (NHERF-1) gene was significantly overexpressed in the invading rim of the tumor specimens when compared to matched, more proliferative core regions (Kislin 2009). In this study, we demonstrated that NHERF-1 functions as a critical “switch” for GBM cells in the differential adoption of a migratory versus proliferative phenotype, potentially regulating the EGFR signaling pathway.

Synergistic Toxicity of Depleted Uranium and UVB Light in Human Skin Cells

Bret Clawson. Northern Arizona University

The flea Oropsylla hirsuta is a known vector of plague among prairie dogs (Cynomys spp.) in the western United States. Plague was introduced to San Francisco in 1900, and spread to ultimately occupy the entire western United States. As O. hirsuta is involved in the regional maintenance of plague, it is valuable to fill gaps of knowledge about the ecological role and gene flow patterns in this flea vector. To support this research, we generated the first partial genome sequence available for O.hirsuta. Using this sequence we isolated and characterized 9 unique microsatellite loci, which contained polymorphic dinucleotide repeats. Using these variable repeat regions as DNA fingerprints we can discern between individuals (as in human forensics) and also gain information about populations. We will use these markers to determine the genetic population structure of O. hirsuta populations. Investigating flea population structure will provide dispersal information that complements other research on the spread of plague in the United States.

Engineering CSS-coated PCL electrospun scaffolds for antibody immobilization and cell capture

Celine Cohn, University of Arizona

Engineered tissue scaffolds can prove more effective if they are able to capture specific cell types. One method by which to engineer tissue scaffolds is electrospinning. Electrospinning Polycaprolactone (PCL) has become a popular choice for scaffold development as it is low-cost and FDA approved. Ideally, electrospun PCL scaffolds would be able to promote the capture of specific cell types. One possible solution is to coat the electrospun PCL with a novel material, cholesterol-succinyl silane (CSS). CSS is a highly stable, polymerizable material that allows for the functional immobilization of antibodies. Three scaffolds were produced: untreated PCL, plasma treated PCL, and PCL with a CSS coating. After scaffold synthesis, it was attempted to functionally immobilize Anti-CD20 and capture Granta-22 cells. Both plasma-treated and CSS-coated PCL scaffolds were able to functionally immobilize antibodies and capture Granta-22 cells.

A Statistical Package for the Biogeographical Analysis of Microbial Communities from “Next Generation” Sequencing Data

Damien Coy and Logan Knecht, Northern Arizona University

With the cost of DNA sequencing data rapidly decreasing, microbial ecology studies are increasing in scope and complexity; more samples are being taken across varying environmental gradients such as time, space (biogeography), pH, or temperature, making analysis even more difficult than in the past. Â Many gradient-analysis statistical methods have been used in traditional macro-scale ecology for years to address these types of data, however, they have not been properly evaluated on micro-scale ecological data to verify that biologically-meaningful results are obtained. To this end, we are developing MiCOS (Microbial Community Observation Statistics), a software package containing statistical tools for the analysis of microbial communities, particularly as applied to the analysis of community composition across gradients. Consolidating many statistical analyses in a single package removes the considerable barrier of moving data between different software packages, which may require different file formats or run only on different operating systems, thereby facilitating a direct comparison of these tools on positive and negative control data. We will present preliminary findings on our analysis of approximately twenty statistical methods on real and simulated test data. The MiCOS software package is open source and will soon be integrated into the popular Quantitiative Insights Into Microbial Ecology (QIIME) software package, making these methods easily accessible to the community as a whole.

The Dynamic Human Microbiome in College Students from Three Campuses

Daniel Domogala, Northern Arizona University

Studies of the human microbiome to date have looked at the microbial ecology across many body sites or time points from a few individuals, or a few body sites from many individuals at a single time point. Here we present preliminary data (the first time point) from a large-scale study of microbial communities from five different body sites of 106 college students at three universities across 10 weekly time points. These data are compiled with rich metadata, obtained through an initial 50 question questionnaire, to support an investigation into factors which may affect the human microbiome over time. An additional questionnaire is completed on a weekly basis to identify any changes in health or lifestyle that may have affected a student’s microbiome in the preceding week. Students began sampling themselves on a weekly basis at five different body sites (forehead, tongue, palm of dominant hand, gut, and armpit) in January/February of 2012. Ten weeks of samples have been collected, and partial 16S rRNA gene sequences have been obtained from the first time point of 106 individuals using an Illumina HiSeq2000 instrument to a median depth of 54,352 sequences/sample. Sequencing targeted the V4/V5 region of the 16S rRNA gene (515F/806R), and data analysis was performed using QIIME 1.4.0-dev. These initial data confirm previous results showing that community composition is primarily determined by body habitat, with gut, tongue, and skin samples clustering independently of one another in UniFrac PCoA plots. Other observations suggest differences correlated with gender and lifestyle choices. Â Including data from additional time points will help identify broader trends in the data.

Assessing Protein Folding in the Presence of Various Crowding Agents

Amber Enriquez, Northern Arizona University

There is a growing appreciation of the role of non-specific macromolecular interactions in cellular processes. Their impact has been difficult to assess for protein folding due to high background signals given off from bulky agents. This study sheds light on the importance of non-specific interactions during folding using a variety of crowding agent solutions including Lysozyme, Ovalbumin, and Bovine Serum Albumin (BSA) as bulk proteins and the polymers glycerol, Ficoll 70, and sucrose to mimic cellular conditions. There has not been a carefully controlled study comparing the impact of all of these. We assessed the folding of Î²-galactosidase in the presence of various crowding solutions to determine the impact of different conditions on refolding yields. Results have shown that protein concentrations of Ovalbumin and BSA at 27.5mg/mL with glycerol enhance refolding while solutions containing only polymers have a minimal impact on refolding yields.

Characterizing the Mechanical Properties of Human Tropoelastin Scaffolds

Audrey Ford, Northern Arizona University

Any material that interfaces with living tissue is considered a biomaterial. In order to be effective, biomaterials, especially biomaterials that are implanted into living tissue must be biocompatible. Biocompatibility is routinely characterized by immune and inflammatory responses by the body, but mechanical properties, like strength and elasticity, of the material are equally important to the function of the material within the surrounding tissues. Human tropoelastin is a versatile and novel biomaterial that has broad potential application as an implantable biomaterial. Tropoelastin can be manipulated by blending it with other proteins or by varying the degree of crosslinking or the type of cross linker used on the scaffold.Â This study aims to characterize the mechanical properties of a range of tropoelastin scaffolds to develop a comprehensive knowledge of its mechanical potential. Understanding the mechanical spectrum of tropoelastin allows it to be used as a customizable biomaterial.

Distribution of Natural Killer Cells in Experimental Autoimmune Encephalomyelitis (EAE)

Soha Ghanian, St. Joseph’s Hospital and Medical Center, Barrow Neurological Institute (High School Student)

Multiple sclerosis (MS) is a chronic, often disabling disease that attacks the central nervous system (CNS), which is composed of the brain, spinal cord, and optic nerves. MS is currently believed to be an immune-mediated disorder caused by the patient’s own immune cells gaining entry into CNS via the impaired blood-brain barrier. This leads to demyelination and scarring of CNS tissue. Natural killer (NK) cells are implicated in the pathogenesis of MS, but their role remains undefined. By using a unique NK1.1-td-Tomato transgenic mouse, the physical infiltration of NK cells in the CNS of EAE, a mouse model of MS, was observed. This study provides support for a potential role of NK cells in the pathogenesis of MS, since NK1.1-td-Tomato is a good model for the study of NK cells in MS.

Electrospun Coaxial Polyvinyl Alcohol – Gelatin Nanofibers: Fabrication & Material Properties

Valerie Gyurko, University of Arizona

Electrospinning has been used to fabricate nanofibrous scaffolds from a variety of synthetic and natural materials, including polyvinyl alcohol (PVA) and gelatin. Although PVA possesses appealing mechanical properties for tissue engineering applications, it lacks adequate cellular recognition sites, thereby limiting it’s bioactivity. In contrast, gelatin has desirable bioactivity but lacks adequate mechanical properties and is difficult to handle. We employed coaxial electrospinning to create nanofibers from these materials in a core/shell structure with gelatin forming the shell and PVA forming the core of the fibers. In this study, scanning electron microscopy (SEM), transmission electron microscopy (TEM), mechanical testing, and cellular studies were utilized to evaluate the morphology and material properties of these fibers. This study yields insight into the potential of combining synthetic and natural polymers together in the engineering composite nanofibers.

All-Trans Retinoic Acid Cytoprotection Against Oxidative Stress

Yurika Isoe, BIO5 Institute Keep Engaging Youth in Science Internship (KEYS)(High School Student)

It has been hypothesized that all-tran acid (ATRA) has cytoprotective properties in scute kidney injury (AKI). Two stable cell lines PK-1 (porcine kidney) and HK-2 (human kidney) were pre-treated with ATRA for 24 hours. Subsequently, hydrogen peroxide (H2O2) was administered to the cells for 3 or 4 hours to induce oxidative stress. Cell viability was assessed by an MTT assay as a measurement of mitochondrial dehydrogenase activity. Both cell lines showed cytoprotection ranging from 10~40%. These results indicate that H2O2 induced oxidative cytotoxicity is mitigated by ATRA cytoprotection.

The Use of Lanthanide-Doped Nanoparticles for Formation of a Temperature Sensitive Coating on Glass

Gihan Joshua, University of Arizona, Biomedical Engineering Graduate Program

The objective of this study was to create a thin film of NaYF4:Er3+/Yb3+ nanoparticles on a glass surface, and observe temperature changes on this glass surface by monitoring temperature dependent emission bands from the nanoparticles. The nanoparticle samples were prepared via thermal decomposition of lanthanide trifluoroacetate precursors at 300oC, and underwent a simple acid treatment process to remove the oleate ligands on their surfaces. Finally the samples were introduced to glass slides that had been functionalized with carboxyl groups, which facilitated binding of the oleate-free nanoparticles to the glass. The nanoparticles on the slides were excited using a 980nm diode laser, and the resulting red and green emission bands were captured using respective filters and a CCD camera. Images were taken at 5oC intervals in the range from 20-65oC, and were used to construct the green-to-red emission ratio, which showed a linear trend with respect to varying temperature.

The Embryonic Development of the Ascidian Notochord: A Simplified Theory

Russell Latterman, Northern Arizona University

The formation of an elongated cell-structure, called a notochord (primitive spinal chord), during the early embryonic development of the ascidian sea creature has been studied by many biologists and mathematicians in hopes of gaining insight into how more-complex organisms develop. How is it that a fairly random clump of dividing cells can suddenly begin to orient themselves into elongated form while also changing shape? We present a computational simulation to support our simpler, and very-arguably more theoretically-sound explanation, than any of the previous models can viably support. This very significant result may contribute to the way mathematicians and biologists form interdisciplinary theories regarding more advanced biological systems. We construct and implement a stochastic model of convergent extension, using a minimal set of assumptions on cell behavior. In addition to the basic assumptions of volume conservation, random cell motion, and cell-cell and cell-ECM adhesion, and a non-standard assumption that cytoskeletal polymerization generates an internal pressure tending to keep cells convex, we find that we need only two conditions for convergent extension. (1) Each cell type has a particular aspect ratio towards which it regulates its geometry. We do not require that cells align in a specific orientation, e.g., to be oriented mediolaterally. (2) The elongating tissue is composed of cells that prefer to be elongated, and these cells must be accompanied by cells which prefer to be round. The latter effectively provide a boundary to capture. In simulations, our model tissue extends and converges to a stacked arrangement of elongated cells one cell wide, an arrangement which is seen in ascidian notochords, but which has not been observed in other models. This arrangement is achieved without any direct mediolateral bias other than that which is provided by the physical edge of the adjacent tissue.

The Effects of Toxoplasma gondii Infection on Mucin Expression in Mouse Intestinal Epithelial Cells

Joshua Mayoral, Northern Arizona University

Toxoplasma gondii is a ubiquitous protozoan parasite, estimated to infect about one-third of the global population of humans. T. gondii infection in hosts is initiated in the intestines, which is lined with a layer of mucus designed to prevent microbial infiltration. Composing this layer of mucus are proteins called mucins. The purpose of this study is to determine whether T. gondii infection causes significant changes in the genetic expression of various mucins located in the intestines; a relationship that previously has not been studied.Â It is hypothesized that T. gondii will cause significant up-regulation of mucin proteins during infection in mouse intestinal cells. To test this hypothesis, MODE-K cells will be cultured and infected with the parasite, after which the genetic material of the cells will be analyzed. The relevance of this study is primarily medicinal. New therapeutic methods may be developed if a link is found between mucin expression and T. gondii infection.

Digital Holography for Biological Imaging

Kristen Milligan, Khalid Wabli and Luke Contreras, Northern Arizona University

We report a technique for obtaining quantitative three dimensional refractive index measurements of biological cells with nanometer precision using digital holography. Digital holographic reconstruction procedures have been implemented on samples such as cancer cells and blood cells that have allowed for the 3D rendering of the sample. Such procedures include an autofocusing program based on a focus metric designed to fit the characteristics of our pure-phase samples, as well as aberration corrections due to different optical components through the application of Zernike polynomials. Our results provide quantitative information about changes in refractive index or morphology of the cell that could be caused by biological processes, disease progression, or drug delivery.

RNF216 (TRIAD3) is a candidate tumor suppressor in advanced tumors

Christopher Murray, Translational Genomics Research Institute

RNF216 (RING finger protein 216) is a gene that maps to chromosome 7p and encodes TRIAD3, which suppresses NF-ÎºB activation. Using next-generation sequencing, two missense mutations were identified in RNF216 in two advanced chemoresistant tumors; one pancreatic and one esthesioneuroblastoma. These mutations were validated using Sanger sequencing. We hypothesize that RNF216 is a tumor suppressor and that mutations lead to TRIAD3’s loss of ability to regulate NF-ÎºB activation. This is highly significant as upregulation of NF-ÎºB activation promotes increased cell survival and proliferation. More importantly, it has been shown that myeloma tumors that have activation of NF-kB are sensitive to the drug bortezomib. In order to test this hypothesis, we have devised a series of functional and pharmacogenomic assays to investigate the consequences of RNF216 loss in T47D and MDA-MB-231 cells, and to determine if loss of TRIAD3 will sensitize cells to bortezomib.

3D Model of Ovarian Carcinoma Microenvironment for Interrogating Intercellular Communication

Gabriel Orsinger, University of Arizona, Biomedical Engineering

With the ultimate goal of designing sophisticated cancer therapies and diagnostics through better understanding the complex tumor microenvironment, we have developed a three-dimensional tissue construct consisting of ovarian surface epithelium and stromal architecture. This 3D model provides co-culture of multiple cell types in a deliberate volumetric arrangement, facilitating a more physiologically relevant microenvironment than standard 2D cell culture for interrogating intercellular communication pathways involved in initiation, progression and metastasis of ovarian carcinomas. In combination with gold-coated plasmon resonant liposomes, this tissue mimic serves as a powerful tool for initiating and monitoring signaling events between and within cancer cell types and for studying pharmocodynamics of liposome-encapsulated and subsequent release of anticancer therapeutics.

A Comparison of Two Common Polycaprolactone-based Biomaterials

P. Daniel Warren, University of Arizona

Recently there has been a significant increase in publications regarding the use of polycaprolactone as a resorbable biomaterial due to its advantages over other aliphatic polymers. This renewed interest has resulted in combinations of PCL with other materials (e.g. polyurethane) to make it more suitable for other applications. To this end, two main trends seem to dominate the literature. One such process simply combines the PCL and another material in solvent to make a blend of the two materials. The other process requires slightly more work to actually covalently link PCL to another material to produce a copolymer. In this work, a common blend of PCL and a commercially available linear polyurethane were compared to a poly(ester-urethane-urea) copolymer based on the amino acid lysine. The materials were characterized to better grasp the impact on properties by the material constituents, to best direct future research on fabricating biomaterials for cardiovascular implant devices.

Using Contact Angle and Wettability to Determine Biocompatibility of Biomedical Implant Devices

Jeff Watson, Northern Arizona University

Biomedical implants are used throughout the body to treat various etiologies or replace damaged or compromised tissues or organs. Extensive biocompatibility testing must be performed prior to their clinical use. Biocompatibility is defined as the “ability of a material to perform with an appropriate response in a specific application” (DF Williams, 1987). A major goal of the current study was to determine if a relationship exists between biocompatibility and wettability of biomaterials. It was expected that a biomaterial with a smaller contact angle (e.g. more hydrophilic) will be more biocompatible in an in vivo animal model, and therefore will be a more optimal biomaterial. Contact angle was measured using a digital camera, high resolution optics, and computer software. Contact angle measurements were compared to in vivo biocompatibility data in efforts to establish a relationship between these two parameters.

Synergistic Toxicity of Depleted Uranium and UVB Light in Human Skin Cells

Mary Zuniga, Northern Arizona University

Most research addresses uranium-induced cancer through its radiological modes of action, ignoring its chemical reactivity. Lungs and kidneys are known target organs for uranium toxicity, but its effects in other organs or tissues are less well understood. It is known that uranium(VI) is photoactivated by UVB-light to form uranium(V) and reactive oxygen species. We therefore hypothesize that combined exposure to uranium and UVB-light may be more toxic and mutagenic to skin than individual exposures. We are testing this hypothesis by exposing cultured human keratinocyte skin cells to depleted uranium (DU)in the presence and absence of UVB-light. We are measuring the effect of these treatments on cell survival, cell damage, and mutation frequency. Preliminary results show increased cytotoxicity when UVB and DU are combined. Further results from this work may provide data to expand the focus of uranium toxicity and include skin as a target organ for its carcinogenic effects.