<p>Obstructive lung diseases limit airflow and gas exchange and have a major impact on a patient’s long-term health. Asthma and chronic obstructive pulmonary disease (COPD) are the most prevalent obstructive lung diseases and represent a major burden on healthcare systems worldwide. It is now accepted that the pathologies associated with these diseases are heterogeneous in nature, and as the function of the lung is determined by its three-dimensional structure, methods to volumetrically evaluate the lung are important tools in furthering the study of these pathologies.</p> <p>Three-dimensional imaging methodologies, such as computed tomography (CT) and single photon emission computed tomography (SPECT), are used clinically in the diagnosis of lung disease, but results are not commonly quantified. In addition, asthma and COPD develop slowly over time and diagnosis normally takes place after the underlying pathologies are well established. Experimental models in small animals, such as rats and mice, allow for the study of disease pathogenesis in a controlled setting and development of quantitative imaging practices for these models provides translational tools for relating results back to the clinic.</p> <p>In this thesis, CT densitometry and ventilation/perfusion (V/Q) SPECT are explored as methods to investigate models of asthma and COPD. CT densitometry is shown to be capable of quantifying allergic inflammation in an asthma model but is of less use in a model of COPD, predominantly due to the relative amounts of inflammation present. However, V/Q imaging is shown to be quite sensitive to the effects of cigarette smoke in a model of COPD and has been used to better understand how pathologies associated with COPD contribute to gas exchange limitation in the lung.</p> <p>The models, imaging techniques, and analysis methods described in this work provide insight into chronic obstructive lung disease and allow for future investigations into how pathologies effect gas exchange. Further, the characterization of the models described in this thesis allows for drug efficacy studies to be performed, both on established and novel treatments. Future research into asthma and COPD will benefit further from the use of threedimensional imaging methodologies because they provide volumetric information on structure and function and can act as a translational bridge between clinical disease and preclinical animal models.</p> / Doctor of Philosophy (Medical Science)

Diabetes is a major risk factor for the development of cataract [3,14,20,22]. In this thesis, we create a model that allows us to understand the incidence of one disease in the context of another; in particular, cataract in the presence of diabetes. The World Health Organization's Vision 2020 blindness-prevention initiative administers surgeries to remove cataracts, the leading cause of blindness worldwide [24]. One of the geographic areas most impacted by cataract-related blindness is Sub-Saharan Africa. In order to plan the number of surgeries to administer, the World Health Organization uses data on cataract prevalence. However, an estimation of the incidence of cataract is more useful than prevalence data for the purpose of resource planning. In 2012, Dray and Williams developed a method for estimating incidence based on prevalence data [5]. Incidence estimates can be further refined by considering associated risk factors such as diabetes. We therefore extend the Dray and Williams model to include diabetes prevalence when calculating cataract incidence estimates. We explore two possible approaches to our model construction, one a detailed extension, and the other, a simplification of that extension. We provide a discussion comparing the two approaches.

No description available.

<p>Food allergies are immune system-driven diseases that lead to reproducible adverse reactions which can be fatal. These severe systemic reactions are primary mediated by immunoglobulin E (IgE) that is derived from B cells which have been activated by T helper type 2 (Th2) cells. While much work has advanced the clinical and pharmacological management of patients with allergic diseases, much remains to be elucidated about how individuals initially acquire allergy. This Thesis details a mechanism linking initial gastrointestinal exposure to peanut (PN) allergen, to the generation of Th2 cells: PN allergen activates epithelial cell secretion of interleukin (IL)-33 and eosinophil degranulation of eosinophil peroxidase, which causes CD103+ dendritic cell (DC) activation and migration to mesenteric lymph nodes where DC OX40L engages naïve T cells to secrete IL-4 in an autocrine/paracrine manner to promote and consolidate Th2 cell differentiation. These events are followed by B cell activation and PN-specific IgE production, which sensitizes mast cells to be hypersensitive to PN re-exposure by causing immediate allergic reactions including anaphylaxis. This is later followed by eosinophilic inflammation that is partially mediated by innate lymphoid cells. As food allergy also serves as a unique model to better understand mechanisms of adaptive immunity, especially Th2 immunobiology, both basic science and clinical implications are discussed in this Thesis. Major themes include Th2 and disease heterogeneity, identification of ‘the original source of IL-4’, an unprecedented <em>in vivo </em>requirement for eosinophils in priming adaptive immune responses, and the need to weigh basic science findings against the human disease <em>in natura </em>litmus test. Looking forward, many questions remain to be answered in the field of food allergy research, but the findings of this Thesis may be one step towards the prevention, management or cure of a disease with growing public concern, potentially fatal consequences, and an unmet need in understanding its pathogenesis.</p> / Doctor of Philosophy (Medical Science)

Background & Aims: Despite advances in surgical care and chemotherapeutic regimens, the five-year survival rate for Stage IV Hepatoblastoma (HB), the predominant pediatric liver tumor, remains at 27%. YAP1 and β-Catenin co-activation occurs in 80% of children’s HB; however, a lack of conditional genetic models precludes exploration of tumor maintenance and therapeutic targets. Thus, the clinical need for a targeted therapy remains unmet. Given the predominance of YAP1 and β-catenin activation in children’s tumors, I sought to evaluate YAP1 as a therapeutic target in HB. Approach & Results: Herein, I engineered the first conditional murine model of HB using hydrodynamic injection to deliver transposon plasmids encoding inducible YAP1S127A, constitutive β-CateninDelN90, and a luciferase reporter to murine liver. Tumor regression was evaluated using in vivo bioluminescent imaging, and tumor landscape characterized using RNA sequencing, ATAC sequencing and DNA foot-printing. Here I show that YAP1 withdrawal in mice mediates >90% tumor regression with survival for 230+ days. Mechanistically, YAP1 withdrawal promotes apoptosis in a subset of tumor cells and in remaining cells induces a cell fate switch driving therapeutic differentiation of HB tumors into Ki-67 negative “hbHep cells.” hbHep cells have hepatocyte-like morphology and partially restored mature hepatocyte gene expression. YAP1 withdrawal drives formation of hbHeps by modulating liver differentiation transcription factor (TF) occupancy. Indeed, tumor-derived hbHeps, consistent with their reprogrammed transcriptional landscape, regain partial hepatocyte function and can rescue liver damage in mice. Conclusions: YAP1 withdrawal, without modulation of oncogenic β-Catenin, significantly regresses hepatoblastoma, providing the first in vivo data to support YAP1 as a therapeutic target for HB. Modulating YAP1 expression alone is sufficient to drive long-term regression in hepatoblastoma because it promotes cell death in a subset of tumor cells and modulates transcription factor occupancy to reverse the fate of residual tumor cells to mimic functional hepatocytes.

Indiana University-Purdue University Indianapolis (IUPUI) / Human induced pluripotent stem (iPS) cells have the unique ability to differentiate into 200 or so somatic cell types that make up the adult human being. The use of human iPS cells to study development and disease is a highly exciting and interdependent field that holds great promise in understanding and elucidating mechanisms behind cellular differentiation with future applications in drug screening and cell replacement studies for complex and currently incurable cellular degenerative disorders. The recent advent of iPS cell technology allows for the generation of patient-specific cell lines that enable us to model the progression of a disease phenotype in a human in vitro model. Differentiation of iPS cells toward the affected cell type provides an unlimited source of diseased cells for examination, and to further study the developmental progression of the disease in vitro, also called the “disease-in-a-dish” model. In this study, efforts were undertaken to recapitulate the differentiation of distinct retinal cell affected in two highly prevalent retinal diseases, Usher syndrome and glaucoma. Using a line of Type III Usher Syndrome patient derived iPS cells efforts were undertaken to develop such an approach as an effective in vitro model for studies of Usher Syndrome, the most commonly inherited disorder affecting both vision and hearing. Using existing lines of iPS cells, studies were also aimed at differentiation and characterization of the more complex retinal cell types, retinal ganglion cells (RGCs) and astrocytes, the cell types affected in glaucoma, a severe neurodegenerative disease of the retina leading to eventual irreversible blindness. Using a previously described protocol, the iPS cells were directed to differentiate toward a retinal fate through a step-wise process that proceeds through all of the major stages of neuroretinal development. The differentiation process was monitored for a period of 70 days for the differentiation of retinal cell types and 150 days for astrocyte development. The different stages of differentiation and the individually derived somatic cell types were characterized by the expression of developmentally associated transcription factors specific to each cell type. Further approaches were undertaken to characterize the morphological differences between RGCs and other neuroretinal cell types derived in the process. The results of this study successfully demonstrated that Usher syndrome patient derived iPS cells differentiated to the affected photoreceptors of Usher syndrome along with other mature retinal cell types, chronologically analogous to the development of the cell types in a mature human retina. This study also established a robust method for the in vitro derivation of RGCs and astrocytes from human iPS cells and provided novel methodologies and evidence to characterize these individual somatic cell types. Overall, this study provides a unique insight into the application of human pluripotent stem cell biology by establishing a novel platform for future studies of in vitro disease modeling of the retinal degenerative diseases: Usher syndrome and glaucoma. In downstream applications of this study, the disease relevant cell types derived from human iPS cells can be used as tools to further study disease progression, drug screening and cell replacement strategies.