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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: 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: TARGETING CELLULAR SIGNALING PATHWAYS IN ESTROGEN RECEPTOR POSITIVE BREAST CANCER
Creator: Gentile, Sonia Kumar
Date: 2018
Collection: Electronic Theses & Dissertations
Description: Estrogen receptor positive (ER+) breast cancer is the most common type of breast cancer diagnosed in women. While usually initially responsive to combinations of chemotherapy with hormonal therapies, resistance to current clinically used treatments is becoming more and more frequent. It is vital to continue to study the mechanisms of resistance to endocrine therapies and discover methods for combating this drug resistance to improve patient survival. The mixed lineage kinase (MLK) inhibitor,...
Show moreEstrogen receptor positive (ER+) breast cancer is the most common type of breast cancer diagnosed in women. While usually initially responsive to combinations of chemotherapy with hormonal therapies, resistance to current clinically used treatments is becoming more and more frequent. It is vital to continue to study the mechanisms of resistance to endocrine therapies and discover methods for combating this drug resistance to improve patient survival. The mixed lineage kinase (MLK) inhibitor, CEP-1347, was studied in culture and in pre-clinical models to evaluate its efficacy, alone and in combination with the clinically available selective estrogen receptor downregulator ICI 182,780, in treating endocrine sensitive and resistant ER+ breast cancers. Using cell lines models, its effects on cell viability, cell death, and cell cycle progression were analyzed, and nuclear and cellular morphology throughout mitosis were examined. Studies were expanded to animals to determine the efficacy of CEP-1347 against tumor growth in a pre-clinical setting. Tumor cell growth and death were studied, as well as the potential for tumor regrowth after the cessation of treatment. Investigation into drug efficacy were expanded into patient derived xenograft (PDX) lines and effects on cell cycle progression were analyzed.The data demonstrate that CEP-1347, especially in combination with ICI 182,780 has the potential to treat endocrine resistant disease through causing a cell cycle arrest which leads to a combination of decreasing proliferation and inducing apoptosis of tumor cells. Furthermore, inhibition of regrowth after cessation of treatment in endocrine sensitive cells suggests that perhaps if used as an earlier line therapy, CEP-1347 in combination with ICI 182,780 may slow or prevent the development of resistance.
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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: 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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