Department of Obstetrics and Gynecology Medical Student Works

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    Development of Cancer-Related Protease Assay Using DNA-Encoded Proteomic Probes
    (2015-05) Ivaturi, Samantha; Jetson, Rachael; Krusemark, Casey
    Proteases function by breaking the peptide bonds within proteins that hold amino acids together. This action, proteolysis, is essential for the proper functioning of numerous biological processes. The activity of this vast group of enzymes is held in a delicate balance in normal living systems, however tumor growth and metastasis can develop as a result of irregular protease activity. A number of protease activities are well known to be misrelated in cancerous cells. The objective of the project is to develop DNA-encoded probes to use as a tool for protease assays that will enable low-cost, high-throughput profiling of enzymatic activity in samples by DNA sequence analysis for the early detection, characterization, and prevention of cancer. The encoding of functional proteomic information in DNA sequence promises to have significant benefits over traditional methods, including highly sensitive detection and capabilities for extensive multiplexing. Model proteases used in this experiment will include caspase 3 and prostate specific antigen (PSA).
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    Isolation and Analysis of a Mycobacteriophage Specific to Mycobacterium Smegmatis
    (2015-12-15) Collins, Angela J.; Ghazali, Danish M.; Li, Yi; Ha, Soo; Clase, Kari L.; Hatfull, Graham
    BACKGROUND: Bacteriophages are viruses that kill bacteria. It is estimated that there are roughly 1031 types of bacteriophages in the world, and that every bacterium has an average of 10 bacteriophages that can infect it. A mycobacteriophage is a type of bacteriophage which has a mycobacterial species as its host. There are 30 known clusters, or types, or mycobacteriophages which all contain very distinct genetic structures, but there are still countless mycobacteriophage populations that need to be isolated. Now that microorganisms are becoming resistant to many antibiotics, the study of bacteriophages is increasingly important because they have an incredibly untapped potential to treat resistant bacteria, eradicate bacterial contaminants in food products, promote ulcer healing, control bacterial growth during fermentation, and much more. The goal of this project was to discover new bacteriophages that might contribute to novel medical solutions. MATERIALS AND METHODS: We plated 29 agar plates for 8 different samples. One sample resulted in plaque growth, from which we performed 9 streak tests, underwent a dilution series and filtration to eliminate the contamination, and harvested a High Titer Lysate. After isolating and purifying the phage genomic DNA, we analyzed the sample and quantified the DNA using a Nanodrop spectrophotometer, and we used agarose gel electrophoresis to separate and analyze the DNA fragments. Finally, we used electron mycrocopy to visualize the physical structure and confirm the successful isolation of a single mycobacteriophage population. DISCUSSION: The cloudiness of the plaque produced by the bacteriophage suggests it is a temperate phage, meaning it switchs between two behavior types regarding replication and survival. Analysis of the physical structure of the mycobacteriophage reveals more information about its genomic DNA; its large capsid diameter suggests lengthy indwelling genomic DNAand its relatively long tail suggests a large tape-measure gene. We were unble to analyze the agarose gel electrophoresis results due to time constraints. However, the results of the genomic DNA sequence will hopefully prove that our bacteriophage is a distinct population. CONCLUSION: We successfully captured, isolated, and purified a single bacteriophage species from the local environment of West Lafayette, Indiana. Upon isolating a single bacteriophage species, we then isolated, purified, restricted, and analyzed the genomic DNA of the phage. Finally, we documented our findings on the Bacteriophage Database as FelixElFago (http://phagesdb.org/phages/FelixElFago/) and sent our DNA to the Sequencing Center.
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    Curriculum Intervention: Assessing Need for and Implementation of Sustainability Development in a Global Context within the First-Year Engineering Curriculum at Purdue University
    (2017-05-04) Collins, Angela J.
    BACKGROUND & OBJECTIVE: Globalization is a world-wide phenomenon that is reshaping international relations in tremendous ways and at impossible rates. Due to rapid advancements in communication, transportation, and information technologies, there has been an increasing connectedness of humanity around the globe. With this trans-global integration comes many benefits, but also countless challenges. It is critical that engineering education facilities constantly update and restructure their curriculum to produce engineers who are capable of tackling the world’s greatest global challenges of this globalization era. Therefore, our team proposes a curriculum intervention of the First-Year Engineering Program. We wish to analyze the depth of global competency concepts taught in the course, with a specific focus on sustainability because it is crucial that young engineers develop an understanding of sustainability and perform their specialized tasks with a sustainable vision in mind. METHODS: To ensure high quality development of Purdue engineers is this area, our research team proposes a curriculum intervention involving two parts: (1) An analysis of the global competency and understanding of sustainability of current engineering sophomores who recently completed the First-Year Engineering (FYE) program at Purdue University; (2) An implementation of concepts from a current Purdue course, CE/EEE 355 Engineering Environmental Sustainability, into the FYE curriculum, as well as an implementation of globalization concepts as needed. Furthermore, our team seeks to answer the following research questions: (1) How consistent is the material that is taught across different sections within the FYE program? (2) How much control and influence does each engineering professor have in covering the topics of globalization and sustainability? (3) What hands-on, practical experience and exposure to globalization concepts are the students getting? EXPECTED OUTCOMES: At the conclusion of this research project, our team expects two tangible outcomes: (1) A plan of implementation of sustainability and globalization concepts into the Purdue engineering curriculum; (2) A set of data measurements and specific goals to determine whether the implemented concepts make a difference. Countless research papers stress the importance of evaluating the impact of new initiatives. Thus, our research team seeks statistical evidence that the concepts learned in CE/EEE 355 make a significant impact on the capability of the student, and the implementation of new concepts based off of CE/EEE 355 make a significant difference when implemented into the FYE engineering curriculum. CONCLUSION: In conclusion, the forces of globalization are leading to rapid changes among global dynamics and international relations, and several effects of globalization, such as poor resource utilization, are beginning to pose a threat to humanity. Therefore, engineers must be able to evolve alongside society and must have the skills to tackle the world’s leading problems. Engineers must also understand the importance of sustainable development to ensure a bright future for younger generations to come. As a result, our team suggests a curriculum intervention of the FYE program to better educated Purdue’s young engineers on issues of globalization and sustainability. Purdue is among the top engineering institutions, but to maintain its relevance and influence, there must be a shift in the curriculum to better prepare its graduates to work in this highly-globalized era.
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    Influenza Group 2 HA Stem-Only Nanoparticles Induce Heterotypic Immune Response
    (2017-08) Asdell, Stephanie M.; Moin, Syed M.; Corbett, Kizzmekia; Boyington, Jeffrey; Graham, Barney S.
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    Endocannabinoids Regulate Cerebellar Granule Cell Differentiation
    (2017-09) Essex, Amanda; Black, Kylie; Baygani, Shawyon; Mier, Tristan; Martinez, Ricardo; Mackie, Ken; Kalinovsky, Anna
    The cerebellum plays a crucial role in learning and execution of complex automated behaviors, including fine motor skills, language, and emotional regulation. Cerebellar development continues throughout an extended postnatal period. The most numerous neurons in the cerebellum, as well as the entire brain, are the cerebellar granule cells (GCs), which are generated in a dedicated secondary proliferative zone, the external granule cell layer (EGL), during the first three postnatal weeks in mice, and over a year in humans. The robust expansion of granule cells during early development is responsible for the majority of cerebellar expansion. Morphological and molecular changes that drive GC proliferation and differentiation have been extensively characterized, starting from the developmental studies by Santiago Ramón y Cajal. GC progenitors (GCPs) proliferate in the outer EGL (oEGL). As they are pushed into the inner EGL (iEGL) by the newly generated GCPs, they exit the cell cycle and begin differentiation, first extending bipolar neurites, followed by tangential migration, and eventually radial migration to the inner granule cell layer (IGL), their target territory. Deregulation of GCPs expansion, proliferation to differentiation switch, or the rate of migration could contribute to abnormal cerebellar size and compartmentalization and disrupt cerebellar circuits’ wiring and function. Endocannabinoids (eCBs) have been identified as key players regulating neuron proliferation and migration in the fore- and mid-brain development, however their role in cerebellar development has not yet been explored in detail. Our preliminary results show robust expression of cannabinoid receptor 1 (CB1) in iEGL GCs, concomitant with expression diacylglycerol lipase α (DGLα) a major enzyme required for the synthesis of eCB 2-arachidonoylglycerol (2-AG), in PCs. Furthermore, our preliminary results show that cerebellar size is reduced in CB1 KOs. In this study we investigate the mechanisms through which eCB signaling may regulate GC proliferation and differentiation, focusing on the GCPs cycle length, rate of differentiation and migration.
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    Human Antibody-Dependent Cellular Cytotoxicity-Mediating Antibodies Do Not Recruit Non-Human Primate CD20+ NK Cells
    (2018-04) Asdell, Stephanie M.; Edwards, R. Whitney; Jha, Shalini; Ferrari, Guido
    The antibody-dependent cell-mediated cytotoxicity (ADCC) response represents one of mechanisms through which the immune system destroys tumor or infected cells. During the ADCC response to HIV-1, natural killer (NK) cells are recruited by natural infection- or vaccine-induced antibodies (Abs) bound to the HIV-1 envelope glycoproteins expressed on infected CD4+ T cells via Fc-gamma Receptor IIIA (Fc-R IIIA). The NK cells’ Ab-mediated recognition of the infected cells leads to the release of granzymes and perforin by degranulation, triggering apoptotic signal pathways in the infected cells and ultimately leading to their elimination. We have optimized an assay to investigate the degranulation of non-human primate (NHP) NK cells from five different NHP donors by measuring levels of CD107a, a marker present on the inner membranes of NK lysosomes. Using one NHP and two human monoclonal antibodies (mAbs), we compared degranulation of NK in NHP splenocytes, NHP peripheral blood mononuclear cells (PBMC), and human PBMC. We observed that both NHP and human mAbs recruited NHP and human NK effector cells. Of note, we examined the activity of CD20+ and CD20- NK cells, with the former being NK cells from a unique subset of NHP NK splenocytes. Our analysis suggests that CD20+ cells likely do not play a role in the NHP ADCC response. In conclusion, we can identify the cellular populations responsible for the Ab-mediated killing of HIV-1 infected cells, and we will be able to further analyze their full functional profile at the level of messenger RNA expression, i.e. their transcriptomic profile.
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    WHI3 Regulation of CDK in S. cerevisiae.
    (2018-04-10) Brown, Lucy; Gihana, Gabriel; Lacefield, Soni
    There are many diseases associated with malfunc4ons in the cell cycle. For instance, aneuploidy—when daughter cells have the abnormal number of chromosomes—results from improper cell division. Diseases that arise from chromosomal abnormali4es range from Down Syndrome to Turner Syndrome to Patau Syndrome, all extremely debilita4ng afflic4ons. Another serious consequence of unregulated cell division is the development of cancers. One of the hallmarks of cancer cells is cell prolifera4on, which is a result of unregulated cell division. Studying cell cycle regula4on in yeast, par4cularly budding yeast, Saccharomyces cerevisiae, allows for a beFer understanding of the human cell cycle. Many of the genes studied in my lab are conserved in humans, meaning that those yeast cell proteins also func4on in human cells. Mito4c cell division—the type of division in which a mother cell produces two iden4cal daughter cells—is regulated by a protein complex called cyclin-dependent kinase (CDK). This protein complex has been intensely studied by cell biologists, yet there is s4ll much that is unknown about how it is controlled. CDK—which must be ac4ve in order for cell division to occur—is regulated by a protein in yeast called Swe1. Swe1 inhibits CDK when the cell is perturbed, thereby hal4ng cell division. One way that the cell can be perturbed is by the dele4on of the protein ELM1. When present, ELM1 regulates the cytoskeleton of the cell. However, when ELM1 is deleted, Swe1 is ac4vated, which results in a delayed cell division and irregularly long buds. These long buds are also the result of sep4n perturba4on. When func4oning properly, sep4n proteins form a ring around the bud neck. We recently found that the dele4on of another protein called WHI3 rescues the ELM1 dele4on-induced cell division delay and long buds. By u4lizing fluorescent microscopy, we have been able to visualize cells lacking both WHI3 and ELM1. Our hypothesis is that WHI3 is somehow involved in regula4on of CDK. In order to test this hypothesis, we are working on experiments to see what happens to CDK and other regulators of cell division when ELM1 is deleted, WHI3 is deleted, and when both WHI3 and ELM1 are deleted. I will be conduc4ng other molecular biology experiments to measure the level of CDK ac4vity in both the cytoplasm and the nucleus. My work will help to elucidate another mechanism by which CDK is regulated, which will contribute to our overall understanding of proper progression through the cell cycle.
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    WHI3 Regulation of Cyclin-Dependent Kinase Activity in Saccharomyces cerevisiae
    (2018-05) Brown, Lucy; Gihana, Gabriel; Lacefield, Soni
    There are many diseases associated with malfunctions in the cell cycle. For instance, aneuploidy—when daughter cells have the abnormal number of chromosomes—results from improper cell division. Diseases that arise from chromosomal abnormalities range from Down Syndrome to Turner Syndrome to Patau Syndrome, all extremely debilitating afflictions. Another serious consequence of unregulated cell division is the development of cancers. One of the hallmarks of cancer cells is cell proliferation, which is a result of unregulated cell division. Studying cell cycle regulation in yeast, particularly budding yeast, Saccharomyces cerevisiae, allows for a better understanding of the human cell cycle. Many of the genes studied in my lab are conserved in humans, meaning that those yeast cell proteins also func4on in human cells. Mitotic cell division—the type of division in which a mother cell produces two identical daughter cells—is regulated by a protein complex called cyclin-dependent kinase (CDK). This protein complex has been intensely studied by cell biologists, yet there is s4ll much that is unknown about how it is controlled. CDK—which must be active in order for cell division to occur—is regulated by a protein in yeast called Swe1. Swe1 inhibits CDK when the cell is perturbed, thereby halting cell division. One way that the cell can be perturbed is by the dele4on of the protein ELM1. When present, ELM1 regulates the cytoskeleton of the cell. However, when ELM1 is deleted, Swe1 is activated, which results in a delayed cell division and irregularly long buds. These long buds are also the result of sep4n perturbation. When functioning properly, sep4n proteins form a ring around the bud neck. We recently found that the dele4on of another protein called WHI3 rescues the ELM1 dele4on-induced cell division delay and long buds. By utilizing fluorescent microscopy, we have been able to visualize cells lacking both WHI3 and ELM1. Our hypothesis is that WHI3 is somehow involved in regulation of CDK. In order to test this hypothesis, we are working on experiments to see what happens to CDK and other regulators of cell division when ELM1 is deleted, WHI3 is deleted, and when both WHI3 and ELM1 are deleted. I will be conducting other molecular biology experiments to measure the level of CDK activity in both the cytoplasm and the nucleus. My work will help to elucidate another mechanism by which CDK is regulated, which will contribute to our overall understanding of proper progression through the cell cycle.
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    Is Operative Diagnosis for Aseptic Revision Total Hip Arthroplasty Related to Patient Reported Outcomes?
    (2018-07) Holder, Erik; Ciesielski, Alex; Ziemba-Davis, Mary; Meneghini, R. Michael
    Background and Hypothesis: Component loosening and instability are the leading causes of revision total hip arthroplasty (THA). The purpose of this study was to compare patient-reported outcomes after revision THA based on failure etiology. We hypothesized that outcomes would differ based on reason for revision. Project Methods: 187 consecutive revision THAs performed between 2010 and 2017 were retrospectively reviewed. Prospectively collected preoperative and minimum one-year Hip Disability and Osteoarthritis Outcome Score/HOOS Jr., UCLA Activity Level, WOMAC Index, and patient satisfaction were assessed based on failure etiology. Demographic variables and covariates were accounted for including sex, age, BMI, ASA classification, heart disease, lumbar spine pathology, narcotic use, fibromyalgia, depression, and autoimmune arthritis. Results: Latest UCLA activity level did not differ based on failure etiology (p=0.381). However, the degree of improvement in activity level was higher (p= 0.04) in patients revised for loosening, instability, and infection compared to ALTR and polyethylene wear. HOOS Jr (p=0.949) and WOMAC total (p=0.147) scores did not differ based on failure etiology at latest follow-up, although patients revised for loosening had greater WOMAC improvement compared to all other groups except polyethylene wear (p=0.016). Satisfaction did not vary based on failure etiology (p=0.365), and demographic and covariates were unrelated to outcomes (p³0.165). Conclusion and Potential Impact: We observed that patient-reported outcomes following revision THA vary based on revision reason and activity level improvement is mitigated patients revised for ALTR and poly wear. These findings may help surgeons and patients alike set expectations for recovery following revision THA.
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    Moringa Oleifera, Miracle Tree and Superfood: Antibacterial Evidence and Nutritional Benefits
    (2018-07-15) Miller, Grace J; Embalabala, Rebecca; Bennett, Claire; Buck, Jacob; Russell, Blake; Holder, Erik; Blanchard, Hope; Henry, Laura
    Moringa oleifera is a tropical plant that has high nutritional qualities. It has been used in India and other parts of Asia as a food and medicine for many years. It is now used in feeding programs in developing nations.  Recently, it has been marketed as a “superfood” in the USA. We have showed in our laboratory that the leaf and seed extracts are antimicrobial. The seeds of M. oleifera can also be used for water purification. We have made some observations from the GCMS data on the medicinal qualities of the leaves, seeds and roots. As a member of the order Brassicales, there is potential for crop improvement for drought and cold tolerance. We are interested in the biochemical and molecular studies of this plant. We present here successful callus formation through tissue culture of M. oleifera.  Several combinations of auxins and cytokinins were tested to induce callus formation of leaf and stem explants of M. oleifera The tissues were evaluated for callus formation. Explants were incubated in full light or dark for 4-8 weeks. We have successfully regenerated moringa plants from callus.