Theses and Dissertations from UMD
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New submissions to the thesis/dissertation collections are added automatically as they are received from the Graduate School. Currently, the Graduate School deposits all theses and dissertations from a given semester after the official graduation date. This means that there may be up to a 4 month delay in the appearance of a give thesis/dissertation in DRUM
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Item QUANTITATIVE ANALYSIS OF INTACT PROTEINS AND RNAS CARRIED BY IMMUNOSUPPRESSIVE EXOSOMES(2016) Geis Asteggiante, Lucia Giorgina; Fenselau, Catherine; Chemistry; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Myeloid-derived suppressor cells (MDSC) are immature myeloid cells that accumulate in the tumor microenvironment of most cancer patients. They are a major obstacle to immunotherapy because they suppress both adaptive and innate immune responses. MDSCs collected from tumor-bearing mice release nano-sized vesicles, called exosomes, which carry biologically active molecules and participate in intercellular communication. Exosomes released by MDSC stimulate migration of other MDSC towards the tumor microenvironment and convert macrophages to a tumor-promoting phenotype. Among the proteins identified in MDSC-released exosomes, S100A8 and S100A9 are low-mass, highly abundant, pro-inflammatory mediators already known to contribute directly to the immune suppressive functions of MDSC. The aim of this work was to successfully interrogate the exosomal intact protein cargo using top-down proteomics, a strategy for protein analysis that has not previously been applied to exosomes of any kind. Several protein forms (proteoforms) were fully characterized, which is critical as post-translational modifications regulate protein functions, cellular location and protein interactions. Additionally, since the tumor promoting activity of MDSC is enhanced by inflammation, we focused on evaluating the effect of increased inflammation on the proteoforms relative abundance using current top-down label-free quantitation techniques (peak intensities and peak areas), and comparing them to our recently validated spectral counting approach. Using spectral counting we were able to estimate differences in abundances of both S100A8 and S100A9 proteoforms. Furthermore, it has been previously reported that exosomes can carry micro RNAs and messenger RNAs. In order to investigate if MDSC-derived exosomes also contain RNAs, a collaborative study was carried out entailing the qualitative and quantitative analysis of miRNAs, mRNA and proteins present in MDSC and their exosomes, and evaluate their changes due to heightened inflammation. The MDSC and exosome protein cargo was analysed by bottom-up proteomics in this case, and the RNA cargo by next generation sequencing. A large number of mRNA and miRNA species were found to be carried by MDSC-derived exosomes and, strikingly, their putative functions were associated to MDSC expansion and suppressive function, and cancer development.Item Utilizing herbicide degradation products and artificial sweeteners as stable tracers to examine agricultural and urban nutrient sources within two tributaries of the Chesapeake Bay(2013) Geis Asteggiante, Lucia Giorgina; Torrents, Alba; McConnell, Laura; Civil Engineering; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Eutrophication of the Chesapeake Bay has contributed to a decline in ecosystem health within the watershed. In this study, MESA (a metabolite of metolachlor) and sucralose were proposed as stable tracers that could be used to discriminate nutrient contributions from agricultural versus urban sources. Two estuaries of the Chesapeake Bay were chosen as model systems: the Choptank River (agricultural) and the Anacostia River (urban). Surface water samples were collected and analyzed for herbicides, metabolites, artificial sweeteners and nutrients. Results supported the hypothesis that sucralose is present in waterways influenced by wastewater plants, and MESA was correlated with changes in nitrate concentration in the Choptank indicating agricultural sources. This work provides proof of concept that tracers can be used in the Chesapeake Bay region to distinguish the influence of urban and agricultural nutrient loads and provides a path to better assess restoration efforts and improved allocation of total maximum daily loads.