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Title: Therapeutically targeting autophagy in non-small cell lung cancer
Creator: Yco, Lisette Pangilinan
Date: 2019
Collection: Electronic Theses & Dissertations
Description: Autophagy is a conserved catabolic pathway which sequesters intracellular components in lysosomes to recycle macromolecules for cell maintenance. The role of autophagy in tumor cells is dynamic and depends on many factors including tumor types, tumor stages, and activity of several tumor suppressors and oncogenes. In this thesis, I wanted to improve our understanding of the unique relationship of autophagy with tumor suppressor p53 and oncogenic KRAS in cancer cells, particularly in NSCLC....
Show moreAutophagy is a conserved catabolic pathway which sequesters intracellular components in lysosomes to recycle macromolecules for cell maintenance. The role of autophagy in tumor cells is dynamic and depends on many factors including tumor types, tumor stages, and activity of several tumor suppressors and oncogenes. In this thesis, I wanted to improve our understanding of the unique relationship of autophagy with tumor suppressor p53 and oncogenic KRAS in cancer cells, particularly in NSCLC. First, I demonstrated that stabilized nuclear wild-type p53 through HDM2 inhibition with MK-8242 or nutlin-3a could induce autophagy in tumor cells through transactivation of several autophagy-related genes (DRAM, FOXO3A, SESN2, and MRCKα) and autophagy core genes (ATG4A and ULK1). In addition, I found that inhibiting of KRAS G12C signaling and suppressing mTORC1 activity by selective KRAS G12C inhibitor, ARS-853, could drive autophagy response in KRAS G12C NSCLC cell lines. Since autophagy could also promote survival under stress induced by several anticancer agents, I designed a combination study using newly reported selective ULK1 inhibitor, ULK-101, with ARS-853 in KRAS mutant NSCLC. Autophagy inhibition with ULK-101 dramatically enhanced the ability of selective KRAS G12C inhibitor to impair the viability of KRAS G12C NSCLC. Together, my study provided evidence that autophagy serves as a survival pathway in tumor cells and that future assessment of small molecule that target autophagy core proteins may be potential cancer therapeutic option in p53 wild-type and KRAS G12C NSCLC.
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Title: The roles of C/EBPbeta and c-Jun in transcription of the gene encoding the murine progesterone receptor
Creator: Do, Han Ngoc
Date: 2014
Collection: Electronic Theses & Dissertations
Description: Progesterone (P) and its receptor, the progesterone receptor (PR), are important for mammary gland development. Moreover, P/PR signaling also contributes to mammary tumorigenesis. Thus, studying the mechanism of PR expression is important in breast cancer research. C/EBPB-deficient mice and mice blocked for AP-1 activity show similar defects in mammary gland development as do PRB-deficient mice, especially during pregnancy, suggesting that these transcription factors might act in the same...
Show moreProgesterone (P) and its receptor, the progesterone receptor (PR), are important for mammary gland development. Moreover, P/PR signaling also contributes to mammary tumorigenesis. Thus, studying the mechanism of PR expression is important in breast cancer research. C/EBPB-deficient mice and mice blocked for AP-1 activity show similar defects in mammary gland development as do PRB-deficient mice, especially during pregnancy, suggesting that these transcription factors might act in the same pathway or may regulate overlapping sets of downstream target genes. An overall decrease in PR observed in sexually mature wild type mice fails to occur in C/EBPB-deficient mice, while no alterations in C/EBPB expression are observed in PR-deficient mice. Moreover, AP-1 has been found to regulate PR expression. These observations suggest that C/EBPB and AP-1 act upstream of PR. This leads us to study the possibility that C/EBPB and AP-1 are required for PR expression.We examined whether C/EBPB participated in the transcriptional regulation of PR expression in the mammary gland. Transient co-transfection of a PR promoter-reporter construct with expression vectors that individually express C/EBPB isoforms (LAP1, LAP2, or LIP) into a mouse mammary carcinoma cell line revealed that all C/EBPB isoforms, surprisingly including LIP (the shortest isoform lacking transactivation domains), can transactivate the PR promoter. Importantly, we found that LIP, in particular, robustly synergizes with an AP-1 member, c-Jun, to drive PR transcription. Consistent with significant roles for C/EBPB and c-Jun in PR expression, knockdown experiments showed that endogenous levels of C/EBPB and c-Jun expression were sufficient to drive the PR promoter-reporter. Additionally, overexpression of LIP elevated PR protein expression from the intact endogenous gene encoding PR. Furthermore, in vivo immunofluorescence studies showed that C/EBPB and PRA expression are mutually exclusive in the mammary epithelium, while PRB is only expressed in cells that express C/EBPB. This suggests an important role for C/EBPB in PRB expression during pregnancy. Then, we studied the mechanism by which LIP and c-Jun synergistically activate the PR promoter. We demonstrated in the reporter assay that the integrity of C/EBP- and AP-1-binding sites was required for the respective C/EBPB; and c-Jun activities on the PR promoter. Moreover, we showed in ChIP assay that efficient promoter occupancy of both LIP and c-Jun and their synergistic transactivation of the PR promoter required at least one C/EBP- and one AP-1-binding site. In addition, as indicating in the sequential ChIP assay, C/EBPB and c-Jun simultaneously occupied PR promoter. This leads us to propose a model where the synergy of C/EBPB and c-Jun in transactivation of the PR promoter is dependent on the two factors mutually stabilizing their recruitment to the PR promoter. Collectively, our data suggest a critical role for C/EBPB, particularly LIP, in PRB expression.
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Title: Rat colon epithelial cells : isolation, cultivation, and benzo(a)pyrene metabolism
Creator: Skrypec, Daniel J.
Date: 1981
Collection: Electronic Theses & Dissertations

Title: THE ROLE OF ARID1A IN ENDOMETRIOSIS-RELATED INFERTILITY
Creator: Marquardt, Ryan Michael
Date: 2022
Collection: Electronic Theses & Dissertations
Description: The inner lining of the uterus, the endometrium, is composed of a luminal epithelial cell layer supported by an underlying stroma which contains epithelial gland structures. These distinct cell types coordinate with complex and dynamic molecular crosstalk tightly controlled by ovarian steroid hormones to regulate a healthy menstrual cycle and support the initiation and maintenance of a healthy pregnancy. Endometriosis occurs when endometrium-like tissue forms lesions outside the uterine...
Show moreThe inner lining of the uterus, the endometrium, is composed of a luminal epithelial cell layer supported by an underlying stroma which contains epithelial gland structures. These distinct cell types coordinate with complex and dynamic molecular crosstalk tightly controlled by ovarian steroid hormones to regulate a healthy menstrual cycle and support the initiation and maintenance of a healthy pregnancy. Endometriosis occurs when endometrium-like tissue forms lesions outside the uterine cavity, and this painful disease afflicts about 10% of reproductive-age women, an estimated 176 million worldwide. Up to 50% of these individuals also experience infertility, and many cases cannot be explained by morphological or ovarian defects, which implicates a uterine environment that is non-receptive to embryo implantation. The molecular basis for the correlation between endometriotic lesion presence and a non-receptive endometrium is unclear, but available evidence suggests that dysregulation of epigenetic regulators may play a role. Expression of AT-rich interaction domain 1A (ARID1A), a chromatin remodeling factor, is lost in some endometriotic lesions and markedly reduced in endometrial biopsies from infertile women with endometriosis, but it is essential in the uterus for fertility. This dissertation evaluates the overarching hypothesis that ARID1A loss connects endometriosis and infertility by causing increased lesion development and a non-receptive endometrium. Chapter 1 provides a review of the current literature on the topics of normal ovarian steroid hormone regulation of endometrial function, the dysregulation that occurs in endometriosis with its clinical implications and therapeutic options, and the specific involvement of ARID1A in endometrial pathophysiology. Chapter 2 delineates a critical role for endometrial epithelial ARID1A in uterine gland function for fertility. Chapter 3 reports the need for endometrial epithelial ARID1A to maintain uterine immune homeostasis during early pregnancy. Chapter 4 explores the involvement of endometrial ARID1A loss in a mouse model of endometriosis-related infertility. Chapter 5 describes a method for in vivo photoacoustic imaging of this endometriosis mouse model through the application of nanoparticle labeling. Finally, Chapter 6 summarizes the findings, discusses conclusions from the synthesized data in the context of the current literature, and provides ideas for future studies of related topics. Together, the studies herein make the case that endometrial ARID1A loss contributes to endometriosis-related infertility by exacerbating endometriotic lesion formation and compromising the ability of the endometrium to maintain the gland function and immune homeostasis necessary for the establishment and maintenance of pregnancy. Continued investigation through studies like these is key to understanding endometrial pathophysiology at the molecular level in order to enable development of targeted treatment options for women suffering the devastating effects of endometriosis and related infertility.
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Title: ZIKA VIRUS-INDUCED PREGNANCY LOSS : LESSONS FROM THE MOUSE EMBRYO
Creator: Watts, Jennifer Leticia
Date: 2021
Collection: Electronic Theses & Dissertations
Description: Adults contracting Zika virus (ZIKV) exhibit mild cold-like symptoms, whereas newborn babiesexhibit fetal defects ranging from mild growth retardation to miscarriage. Aside from transmission via mosquito, ZIKV is also sexually transmitted, which introduces the possibility that ZIKV infection could occur shortly after conception. However, the mechanisms underlying ZIKV-induced birth defects in early development are not understood. I hypothesize that sexually transmitted ZIKA virus infects...
Show moreAdults contracting Zika virus (ZIKV) exhibit mild cold-like symptoms, whereas newborn babiesexhibit fetal defects ranging from mild growth retardation to miscarriage. Aside from transmission via mosquito, ZIKV is also sexually transmitted, which introduces the possibility that ZIKV infection could occur shortly after conception. However, the mechanisms underlying ZIKV-induced birth defects in early development are not understood. I hypothesize that sexually transmitted ZIKA virus infects embryos around the time of conception, leading to the most severe congenital defects. Consistent with this hypothesis, I have discovered that candidate proviral factors are present in mouse embryo-derived stem cell lines and preimplantation development. However, embryo-derived stem cell lines exhibited low viral infection and replication. Nevertheless, Puerto Rican (ZIKVPR) and the Ugandan (ZIKVUG) strains of ZIKV caused two-cell embryos to undergo developmental arrest. Moreover, infected blastocyst exhibited reduced SOX2 expression, an epiblast cell marker, CDX2 a trophectoderm cell marker, and SOX17, a primitive endoderm marker. Therefore, my results suggest that preimplantation ZIKV infection causes embryonic demise or embryonic cell fate defects depending on the time of infection. My studies are significant to human health because they will further our knowledge of viral infection in early pregnancy and the outcomes.
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Title: EXPRESSION AND ROLES OF BLASTOCYST LINEAGE-DETERMING GENES DURING SOMATIC CELL REPROGRAMMING
Creator: Moauro, Alexandra
Date: 2022
Collection: Electronic Theses & Dissertations
Description: In order to properly use stem cells, it is important that we first understand how these cells are establish and maintained. One of the most widely used stem cells are induced pluripotent stem cells (iPSCs) which provide great therapeutic promise and a novel source of ethical stem cells for research models. iPSCs are created by overexpression Oct4, Sox2, Klf4 and c-Myc (OSKM) in a somatic cell. As studies have sought to improve reprogramming efficiency and develop the most embryonically...
Show moreIn order to properly use stem cells, it is important that we first understand how these cells are establish and maintained. One of the most widely used stem cells are induced pluripotent stem cells (iPSCs) which provide great therapeutic promise and a novel source of ethical stem cells for research models. iPSCs are created by overexpression Oct4, Sox2, Klf4 and c-Myc (OSKM) in a somatic cell. As studies have sought to improve reprogramming efficiency and develop the most embryonically identical stem cells, our lab has uncovered that OSKM is not a specific cocktail for pluripotency formation. Instead OSKM induces additional cell fates including the formation of a multipotent stem cell termed induced extraembryonic endoderm stem (iXEN) cells. This raises the question as to how two distinct stem cell types arise in parallel. Interestingly, in embryo development we observe the same pluripotent and multipotent extraembryonic endoderm lineages form in parallel. Using our knowledge of normal embryo development, I set out to identify what blastocyst lineage markers can help us identify early iPSC and iXEN colonies as they start to form and mature. Of these markers, we observed that endogenous OCT4 is expressed in both iXEN and iPSC colonies. Based on the expression pattern of the key embryonic transcription factor, OCT4, we further focused on how this transcription factor may have a dual role in establishing iPSC and iXEN fates. Lastly, we altered the reprogramming cocktail using additional embryonic transcription factors to determine how these factors affect the propensity for pluripotency or extraembryonic endoderm fate.
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Title: Impact of ploidy on morphological variation in Arizona phlox, Phlox amabilis (Polemoniaceae)
Creator: Chansler, Matthew Thomas
Date: 2015
Collection: Electronic Theses & Dissertations
Description: Polyploidy is an important factor in the evolution and ecology of flowering plants. A better understanding of the kind and degree of morphological differentiation among ploidy levels within a species can help explain further how polyploidy affects biodiversity. How widespread is the impact of ploidy across the phenotype of a species? Which aspects of morphology vary, and do they vary consistently? How does ploidy relate to overall morphological diversity? Do ploidy levels have detectable...
Show morePolyploidy is an important factor in the evolution and ecology of flowering plants. A better understanding of the kind and degree of morphological differentiation among ploidy levels within a species can help explain further how polyploidy affects biodiversity. How widespread is the impact of ploidy across the phenotype of a species? Which aspects of morphology vary, and do they vary consistently? How does ploidy relate to overall morphological diversity? Do ploidy levels have detectable phenotypic profiles? Finally, are there morphological differences between populations, potentially due to environment or evolutionary changes since formation apparent in natural populations? I assessed morphological variation within Arizona phlox, Phlox amabilis. This species of conservation concern is endemic to Arizona, and prior work has detected diploid, tetraploid, and hexaploid populations. I sampled 11 populations of P. amabilis, covering a large portion of the species’ range. A wide array of morphological features, including characters that described cell size, overall habit, leaf dimensions, and floral dimensions, were measured for up to 25 plants at each population. Significant differences were detected in 15 out of 27 characters using mixed GLM. A large amount of overall morphological variation is explained by the differences between ploidy levels, and each ploidy level can be described by a specific multivariate phenotype with 95% accuracy. Finally, although overall structuring was influenced by ploidy, differences among populations still contributed a high degree of variation in the morphospace of Phlox amabilis. This morphological assessment will be integrated with ecological and genetic data to build a more complete understanding of the interplay between these factors and ploidy in Phlox amabilis.
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Title: 2B4 IS A CHECKPOINT MOLECULE FOR iNKT CELL ANTI-TUMOR RESPONSE
Creator: Bahal, Devika Naresh
Date: 2022
Collection: Electronic Theses & Dissertations
Description: Invariant natural killer T (iNKT) cells are robust cytotoxic effectors and immune modulators, which makes them ideal candidates for cancer immunotherapy. However, the use of iNKTs for cellular therapy against cancer has been limited due to their transient response in pre-clinical trials. Although TCR-CD1d interactions are generally required for iNKT cell cytotoxicity, the receptors and signaling mechanisms that co-operate with the TCR to promote maximal anti-tumor responses are poorly...
Show moreInvariant natural killer T (iNKT) cells are robust cytotoxic effectors and immune modulators, which makes them ideal candidates for cancer immunotherapy. However, the use of iNKTs for cellular therapy against cancer has been limited due to their transient response in pre-clinical trials. Although TCR-CD1d interactions are generally required for iNKT cell cytotoxicity, the receptors and signaling mechanisms that co-operate with the TCR to promote maximal anti-tumor responses are poorly understood. Therefore, elucidating the mechanisms that regulate anti-tumor responses is critical for the development of effective iNKT-based therapies. Our efforts have shown that 2B4, a SLAM receptor, when expressed on iNKTs reduces their cytotoxic response against lymphoma cells. Surprisingly, 2B4 is not expressed on resting iNKTs but gets rapidly upregulated via stimulation through the TCR. 2B4 has two isoforms, which are splice variants of each other, of which the inhibitory long form is predominantly expressed in activated iNKTs. Our data show that 2B4 is a checkpoint molecule and has an inhibitory role in iNKT cell cytotoxicity. Indeed, when we overexpressed 2B4 in an iNKT cell hybridoma, the killing capacity of the iNKT cell line was abrogated. Moreover, 2B4 can be converted to a potent activating receptor by swapping its intracellular domains with proline motifs, which drastically augments tumor cell lysis. Taken together, this study highlights the important role of 2B4 in iNKT cell cytolysis and broadens the knowledge of immunoregulatory receptors in iNKT cells for future applications in cancer therapy.
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Title: The cytologic responses of normal beagle dogs utilizing the skin window technic
Creator: Drees, David T.
Date: 1966
Collection: Electronic Theses & Dissertations

Title: Nanoengineered tissue scaffolds for regenerative medicine in neural cell systems
Creator: Tiryaki, Volkan Mujdat
Date: 2013
Collection: Electronic Theses & Dissertations
Description: Central nervous system (CNS) injuries present one of the most challenging problems. Regeneration in the mammal CNS is often limited because the injured axons cannot regenerate beyond the lesion. Implantation of a scaffolding material is one of the possible approaches to this problem. Recent implantations by our collaborative research group using electrospun polyamide nanofibrillar scaffolds have shown promising results in vitro and in vivo. The physical properties of the tissue scaffolds have...
Show moreCentral nervous system (CNS) injuries present one of the most challenging problems. Regeneration in the mammal CNS is often limited because the injured axons cannot regenerate beyond the lesion. Implantation of a scaffolding material is one of the possible approaches to this problem. Recent implantations by our collaborative research group using electrospun polyamide nanofibrillar scaffolds have shown promising results in vitro and in vivo. The physical properties of the tissue scaffolds have been neglected for many years, and it has only recently been recognized that significant aspects include nanophysical properties such as nanopatterning, surface roughness, local elasticity, surface polarity, surface charge, and growth factor presentation as well as the better-known biochemical cues.The properties of: surface polarity, surface roughness, local elasticity and local work of adhesion were investigated in this thesis. The physical and nanophysical properties of the cell culture environments were evaluated using contact angle and atomic force microscopy (AFM) measurements. A new capability, scanning probe recognition microscopy (SPRM), was also used to characterize the surface roughness of nanofibrillar scaffolds. The corresponding morphological and protein expression responses of rat model cerebral cortical astrocytes to the polyamide nanofibrillar scaffolds versus comparative culture surfaces were investigated by AFM and immunocytochemistry. Astrocyte morphological responses were imaged using AFM and phalloidin staining for F-actin. Activation of the corresponding Rho GTPase regulators was investigated using immunolabeling with Cdc42, Rac1, and RhoA. The results supported the hypothesis that the extracellular environment can trigger preferential activation of members of the Rho GTPase family, with demonstrable morphological consequences for cerebral cortical astrocytes. Astrocytes have a special role in the formation of the glial scar in response to traumatic injury. The glial scar biomechanically and biochemically blocks axon regeneration, resulting in paralysis. Astrocytes involved in glial scar formation become reactive, with development of specific morphologies and inhibitory protein expressions. Dibutyryl cyclic adenosine monophosphate (dBcAMP) was used to induce astrocyte reactivity. The directive importance of nanophysical properties for the morphological and protein expression responses of dBcAMP-stimulated cerebral cortical astrocytes was investigated by immunocytochemistry, Western blotting, and AFM. Nanofibrillar scaffold properties were shown to reduce immunoreactivity responses, while PLL Aclar properties were shown to induce responses reminiscent of glial scar formation. Comparison of the responses for dBcAMP-treated reactive-like and untreated astrocytes indicated that the most influential directive nanophysical cues may differ in wound-healing versus untreated situations.Finally, a new cell shape index (CSI) analysis system was developed using volumetric AFM height images of cells cultured on different substrates. The new CSI revealed quantitative cell spreading information not included in the conventional CSI. The system includes a floating feature selection algorithm for cell segmentation that uses a total of 28 different textural features derived from two models: the gray level co-occurance matrix and local statistics texture features. The quantitative morphometry of untreated and dBcAMP-treated cerebral cortical astrocytes was investigated using the new and conventional CSI, and the results showed that quantitative astrocyte spreading and stellation behavior was induced by variations in nanophysical properties.
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Title: Biochemical analysis of transformation-sensitive alterations in the substratum associated material of chicken embryo fibroblasts
Creator: Blenis, John
Date: 1983
Collection: Electronic Theses & Dissertations

Title: THE NUCLEO-CYTOPLASMIC FUNCTION OF ACTIN AND ACTIN DEPOLYMERIZATION FACTORS IN PLANT IMMUNITY
Creator: Li, Pai
Date: 2022
Collection: Electronic Theses & Dissertations
Description: The plant immune system is a multi-phase complex network that involves the collaboration of multiple subcellular structures. In the past two decades, the core signaling pathways of the immune process, including pattern-triggered immunity (PTI), effector-triggered immunity (ETI), and systemic acquired resistance (SAR), as well as the behavior of organelles, have been revealed to a level of clarity that is able to describe a general and well-covered process of the immune response. However,...
Show moreThe plant immune system is a multi-phase complex network that involves the collaboration of multiple subcellular structures. In the past two decades, the core signaling pathways of the immune process, including pattern-triggered immunity (PTI), effector-triggered immunity (ETI), and systemic acquired resistance (SAR), as well as the behavior of organelles, have been revealed to a level of clarity that is able to describe a general and well-covered process of the immune response. However, there are still many events during the immune response that remain mysterious. For instance, while higher plants live a sessile lifestyle, there are countless intracellular motions mediated by the cytoskeleton (including its associated proteins) in response to the external triggers, such as the invasion of pathogens. As our knowledge of plant immunity accumulates, the deficiency in knowledge on how immune signaling regulates the behavior of the cytoskeleton as a critical aspect of defense response, howbeit, becomes more evident. Therefore, this is a field of research that calls for powerful toolboxes to facilitate the analysis of the cytoskeleton in the context of immunity, as well as instructive biological model(s) that guide the direction of the multifarious studies. In this dissertation, I focus on the summary and prospective discussion on the immune function of the actin cytoskeleton and, more importantly, describe my original studies on two major aspects of this topic. First, a prerequisite to functional study of the actin cytoskeleton in the cytoplasm is the ability to accurately describe the status of the cytoskeleton. To achieve this goal, I developed an algorithm, namely implicit Laplacian of enhanced edge (ILEE), to accurately identify and analyze the biological status of the cytoskeleton from confocal image samples. This method significantly improves the accuracy, stability, and robustness of cytoskeleton segmentation, solves other technical hindrances, and enables abundant information to be extracted from images for biological interpretation (see Chapter 2). The ILEE algorithm will further help me to explore the phenotypes of actin architecture in response to immune signaling, which was not previously available due to the lack of the toolbox. Also, the ILEE has been packaged as a library released publicly to benefit the community with a powerful cytoskeleton analysis platform.For the second project of my total research, I focused on the immune function of the actin cytoskeleton in the nucleus. Previously, some Arabidopsis actin depolymerization factors were reported to genetically contribute to plant immunity by unknown mechanism(s), and my story began with a novel activity identified among Arabidopsis actin depolymerization factors – to interact with WRKYs, the stress-responsive transcription factors. During my research, I proved that certain ADFs can form a complex with WRKYs that binds to targeted promoters, hence regulating the activity of WRKYs and playing a positive role in the immune response. The knowledge obtained through this study, in combination with previous research (Lu et al., 2020; Porter et al., 2012a) of my lab, can be summarized into a biological model, in which ADF mediates a nuclear-cytoplasmic immune regulation that systemically facilitates both cytoskeleton dynamics and pro-immune transcriptome reprogramming. In general, this study reveals a novel yet general pattern of cytoskeleton mediated transcriptional regulation, as ADF and perhaps other components of the actin cytoskeleton can shuttle between the cytoplasm and nucleus to form a network with a higher level of complexity. As a potential broader impact, the application range of this model includes but is not necessarily limited to plant immunity.
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Title: Cytology and seed set studies in the pansy, Viola tricolor Hortensis L
Creator: Emino, Everett Raymond, 1942-
Date: 1967
Collection: Electronic Theses & Dissertations

Title: Dietary manipulation of natural killer cell biology through refeeding of previously calorically restricted mice
Creator: Clinthorne, Jonathan F.
Date: 2013
Collection: Electronic Theses & Dissertations
Description: The dynamic role of natural killer (NK) cells in immunology has been demonstrated in numerous fashions, proving they are much more than "natural killers". However, NK cells are perhaps still the least well understood lymphocyte, due to their relatively low abundance and the limited transgenic models available for study. An innate immune cell that plays a critical role in providing early immunity against viral infections and cancers, NK cells are becoming increasingly recognized for shaping...
Show moreThe dynamic role of natural killer (NK) cells in immunology has been demonstrated in numerous fashions, proving they are much more than "natural killers". However, NK cells are perhaps still the least well understood lymphocyte, due to their relatively low abundance and the limited transgenic models available for study. An innate immune cell that plays a critical role in providing early immunity against viral infections and cancers, NK cells are becoming increasingly recognized for shaping and directing immune responses. Accounting for approximately 5-25% of peripheral blood mononuclear cells in humans and 5-10% of lymphocytes in murine circulation, NK cells have very similar functional attributes in both species, making mice an ideal model system for the study of NK cell biology. Studies in mice have revealed the critical importance of NK cells in providing tumor surveillance as well as early protection from viral infections. Preclinical studies have underscored the utility of using NK cells as an immunotherapeutic technique to combat various cancers, while NK cell function is often associated with positive or negative outcomes in various disease states. Furthermore, various lifestyle factors have been found to positively or negatively influence NK cell function. Among these lifestyle factors, diet has gained notoriety as being capable of influencing the homeostasis and function of NK cells. However, the mechanisms by which diet influences NK cells are not fully understood, highlighting the need for a better understanding of the molecular and cellular mechanisms by which diet influences NK cell development. Thus, our laboratory has extensively studied the effects of the restriction of energy intake, or caloric restriction (CR) on NK cell function and biology at the organismal, cellular, and molecular level. Here we describe a series of experiments investigating the role of energy intake on immunity to influenza virus, with a focus on NK cells. We describe a series of studies that identify the specific changes to NK cells induced by CR, both beneficial and potentially damaging. In these experiments we show that CR results in fewer NK cells with a mature phenotype, and that expression of transcription factors critical for NK cell maturation are reduced by CR. We also demonstrate that thymic derived NK cells are present in normal numbers in CR mice and have enhanced function. In a series of experiments demonstrating the intricate relationship between immunity and metabolism we show how refeeding of CR mice restores NK cell homeostasis and function, both before, and during influenza infection. Using in vitro techniques combined with ex vivo analysis of metabolic signaling pathways, we provide potential mechanisms by which CR impairs NK cell maturation. These studies serve to highlight the critical role of optimal nutrition in maintaining NK cell homeostasis and function. To our knowledge, this dissertation is the first data presented that clearly details the effects of CR on NK cell development and homeostasis, as well as the molecular and biochemical pathways mediating this effect.
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Title: SIGNALING MECHANISMS OF PULMONARY ARTERIAL HYPERTENSION
Creator: Ji, Yajing
Date: 2022
Collection: Electronic Theses & Dissertations
Description: Pulmonary arterial hypertension (PAH) is a severe and life-threatening disease that is characterized by elevated pulmonary blood pressure. A challenge in treating PAH is that while the current generation of therapeutics alleviate symptoms, they fail to target the underlying causes of the disease. Initially it was thought that PAH is caused by increased pulmonary vasoconstriction; it is now understood that PAH mainly results from remodeling of the pulmonary vasculature. Further...
Show morePulmonary arterial hypertension (PAH) is a severe and life-threatening disease that is characterized by elevated pulmonary blood pressure. A challenge in treating PAH is that while the current generation of therapeutics alleviate symptoms, they fail to target the underlying causes of the disease. Initially it was thought that PAH is caused by increased pulmonary vasoconstriction; it is now understood that PAH mainly results from remodeling of the pulmonary vasculature. Further characterization of the underlying mechanisms of PAH will identify newpharmacological targets to treat PAH. In this dissertation I seek to address this challenge from three distinct perspectives. In Chapter 2, I investigated the signaling network downstream of TGFβ and highlighted the MRTF/SRF pathway as potential therapeutical targets for PAH given its pivotal role regulating expression of contractile proteins in PASMCs. In Chapter 3, I aim to test whether TGFβ and the silencing of BMPR2, a member of the TGFβ family of receptors, contribute to the activation of lung fibroblasts in vitro. My results presented do not replicate the role of BMPR2 silencing found in other studies. This could be caused by the relatively short duration of BMPR2 silencing in our system. Finally, in Chapter 4, I perform a combined meta-analysis of several publicly available transcriptomic datasets of lung tissues from PAH patients. Using this approach, I identify PAH-associated signaling pathways, and chemical compounds which reverse a PAH-associated gene expression signature. My findings also suggest that while we bin PAH patients into various subtypes in the clinic, on a transcriptional level, PAH patients tend to group into distinct gene expression clusters without relying on their clinical subtype. These findings improve our understanding of PAH biology and also highlight several potential drug targets for PAH.
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Title: Reprogramming to the nervous system : a computational and candidate gene approach
Creator: Alicea, Bradly John
Date: 2013
Collection: Electronic Theses & Dissertations
Description: The creation of stem-like cells, neuronal cells, and skeletal muscle fibers from a generic somatic precursor phenotype has many potential applications. These uses range from cell therapy to disease modeling. The enabling methodology for these applications is known as direct cellular reprogramming. While the biological underpinnings of cellular reprogramming go back to the work of Gurdon and other developmental biologists, the direct approach is a rather recent development. Therefore, our...
Show moreThe creation of stem-like cells, neuronal cells, and skeletal muscle fibers from a generic somatic precursor phenotype has many potential applications. These uses range from cell therapy to disease modeling. The enabling methodology for these applications is known as direct cellular reprogramming. While the biological underpinnings of cellular reprogramming go back to the work of Gurdon and other developmental biologists, the direct approach is a rather recent development. Therefore, our understanding of the reprogramming process is largely based on isolated findings and interesting results. A true synthesis, particularly from a systems perspective, is lacking. In this dissertation, I will attempt to build toward an intellectual synthesis of direct reprogramming by critically examining four types of phenotypic conversion that result in production of nervous system components: induced pluripotency (iPS), induced neuronal (iN), induced skeletal muscle (iSM), and induced cardiomyocyte (iCM). Since potential applications range from tools for basic science to disease modeling and bionic technologies, the need for a common context is essential.This intellectual synthesis will be defined through several research endeavors. The first investigation introduces a set of experiments in which multiple fibroblast cell lines are converted to two terminal phenotypes: iN and iSM. The efficiency and infectability of cells subjected to each reprogramming regimen are then compared both statistically and quantitatively. This set of experiments also resulted in the development of novel analytical methods for measuring reprogramming efficiency and infectability. The second investigation features a critical review and statistical analysis of iPS reprogramming, specifically when compared to indirect reprogramming (SCNT-ES) and related stem-like cells. The third investigation is a review and theoretical synthesis which stakes out new directions in our understanding of the direct reprogramming process, including recent computational modeling endeavors and results from the iPS, iN and induced cardiomyocyte (iCM) experiments. To further unify the outcomes of these studies, additional results related to Chapter 2 and directions for future research will be presented. The additional results will allow for further interpretation and insight into the role of diversity in direct reprogramming. These future directions include both experimental approaches (a technique called mechanism disruption) and computational approaches (preliminary results for an agent-based population-level approximation of direct reprogramming). The insights provided here will hopefully provide a framework for theoretical development and a guide for traditional biologists and systems biologists alike.
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Title: SULFATED AND SIALYLATED N-ACETYL-LACTOSAMINE AS BIOMARKER OF SUBPOPULATIONS OF PANCREATIC DUCTAL ADENOCARCINOMAS
Creator: Hsueh, Peter Yiping
Date: 2019
Collection: Electronic Theses & Dissertations
Description: The sialyl Lewis A (sLeA) glycan forms the basis of the CA19-9 blood test and is the current biomarker for pancreatic ductal adenocarcinoma (PDAC). However, it is not elevated in approximately 25% of PDAC patients and it also has difficulties in diagnosing early-stage PDAC. My overarching goal was focusing on improving precision of overall PDAC diagnostics. I hypothesized that other glycans within the Lewis blood group family besides sLeA are aberrantly increased in the subpopulation of PDAC...
Show moreThe sialyl Lewis A (sLeA) glycan forms the basis of the CA19-9 blood test and is the current biomarker for pancreatic ductal adenocarcinoma (PDAC). However, it is not elevated in approximately 25% of PDAC patients and it also has difficulties in diagnosing early-stage PDAC. My overarching goal was focusing on improving precision of overall PDAC diagnostics. I hypothesized that other glycans within the Lewis blood group family besides sLeA are aberrantly increased in the subpopulation of PDAC patients who do not secret sLeA into their blood. To test the hypothesis, two specific approaches were implemented in this study: 1) Profile an isomer of sLeA, named sialyl-Lewis X (sLeX), and glycans with fucosylated motifs in the plasma of sLeA-low PDAC patients using antibody and lectin microarray method; and 2) Test the sulfated and sialylated glycans derived from type 2 N-acetyl-lactosamine precursor in subpopulations of PDACs using a novel on-chip analysis method.In the first approach, I profiled the levels of multiple glycans and glycosylated mucins in plasma from two cohorts of 200 and 116 test subjects with PDACs and non-malignant disease patients. From these screens, I found significant increases in two categories of glycans: sialyl Lewis X variants, presented both in sulfated and non-sulfated forms, and the sialylated type 1 N-acetyl-lactosamine. These glycans are increased in distinct groups of PDAC patients and contribute to the improved accuracy of a biomarker panel.Thus, I concluded that detecting other glycans within the Lewis blood-group besides sLeA has the potential to improve diagnoses of PDAC patients.To further elucidate the structural nuances of sialyl Lewis X variants from initial screen, I developed a new assay called On-chip Glycan Modification and Probing and a complementary computational algorithm to accurately analyze novel sulfated and sialylated glycans from plasma of pancreatic cancer patients. In detailed structural information, I observed strong evidences of sulfated and sialylated type 2 N-acetyl-lactosamine glycans overexpressed in plasma of PDAC patients and pancreatic cancer cell lines, but not in the plasma of healthy people. In addition, the sulfated and sialylated type 2 N-acetyl-lactosamine glycans presented on a specific mucin, MUC5AC, was statistically associated (p < 0.001) with short time-to-progression of PDAC patients, but CA19-9 test was not. I concluded sulfated and sialylated type 2 N-acetyl-lactosamine glycans presented on MUC5AC were new serological biomarkers that could improve precision of current practices for diagnosis and prognosis of PDACs patients.
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Title: Peroxisome associated proteolytic processes in Arabidopsis
Creator: Kaur, Navneet
Date: 2013
Collection: Electronic Theses & Dissertations
Description: Peroxisomes are small single-membrane-bounded organelles that play key roles in development and metabolism in most eukaryotic organisms. Peroxisome functions encompass ß-oxidation of fatty acids, detoxification of hydrogen peroxide and the metabolism of a range of biochemical compounds including glyoxylate, glycolate, urate, polyamines, benzoate, phylloquinone, bile acids and plasmalogen to name a few, as well as synthesis of plant hormones such as jasmonic acid (JA) and indole-3-butyric acid...
Show morePeroxisomes are small single-membrane-bounded organelles that play key roles in development and metabolism in most eukaryotic organisms. Peroxisome functions encompass ß-oxidation of fatty acids, detoxification of hydrogen peroxide and the metabolism of a range of biochemical compounds including glyoxylate, glycolate, urate, polyamines, benzoate, phylloquinone, bile acids and plasmalogen to name a few, as well as synthesis of plant hormones such as jasmonic acid (JA) and indole-3-butyric acid (IBA). Peroxisomes exhibit great functional diversity largely because of a plastic proteome that varies greatly depending on the environmental condition, tissue type or developmental stage of the specific organism. Proteins in peroxisomes are imported by the actions of conserved machinery that includes several proteins known as peroxins that are important for peroxisome biogenesis. Proteins destined for the peroxisome matrix contain peroxisome targeting signal (PTS) enabling their recognition by cytosolic receptor proteins that transport them to the peroxisome. Although we have an increased understanding of how peroxisomal protein import is accomplished, we know little about how proteins in peroxisomes are degraded. In this research, I provide evidence that RING domains of three peroxisomal membrane proteins AtPEX2, AtPEX10 and AtPEX12 have E3 ligase activity. I further show that AtPEX2 specifically interacts with two homologous ubiquitin receptor proteins, DSK2a and DS2Kb that have been implicated as adapters linking ubiquitination and 26S proteasome-based degradation events. DSK2 amiRNA lines lacked obvious plant growth phenotypes and were not compromised in peroxisome functions, suggesting that functional redundancies exist among ubiquitin receptor proteins. My results indicate that Arabidopsis RING peroxins and DSK2s can together form a peroxisome membrane associated degradation system. I also explored the role of a predicted ovarian tumor-like cysteine protease (OCP1) in Arabidopsis. OCP1 was found to be a novel plant specific peroxisomal protein with a canonical C-terminal PTS1 and a novel N-terminal PTS2. Analysis of mutant lines revealed that OCP1 influences IBA metabolism in the peroxisome. Further, ocp1 mutants show retarded degradation of two transiently expressed seedling peroxisome enzymes, isocitrate lyase (ICL) and malate synthase (MS) suggesting that OCP1 has a role in the timely removal of ICL from seedling peroxisomes. In summary, these studies add significantly to our knowledge of proteolysis in plant peroxisomes and open up several avenues for future investigations that may have ramifications in agriculture and biomedical applications.
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