Life Sciences Commons^™

The Role Of Connexin And Pannexin Large-Pore Channels In Hearing, Julia Abitbol

Electronic Thesis and Dissertation Repository

Connexin and pannexin large-pore channels allow the regulated passage of small molecules at sites of cell-cell contacts, and from the cytosol to the extracellular milieu, respectively. Since it has been known for many years that Cx26 and Cx30 gap junction proteins are crucial in hearing we propose that Cx43 might also be important in hearing. Here we used two different genetically modified mouse lines that contain systemic Cx43 gene mutations that reduces gap junctional intercellular communication (GJIC) to examine whether Cx43 is also important for proper hearing function. Furthermore, since pannexins have also been postulated to be involved in auditory …

The Role Of Xenopus Laevis Reck In Ecm Remodeling And Tissue Patterning, Jessica Willson

Electronic Thesis and Dissertation Repository

Proper cell-cell and cell-extracellular matrix (ECM) interactions are vital for cell migration and patterning of the vertebrate embryo. Matrix metalloproteinases (MMPs) and their inhibitors, reversion-inducing cysteine-rich proteins with Kazal motifs (RECK) and tissue inhibitors of metalloproteinases (TIMPs), are all differentially expressed during embryogenesis to regulate such ECM remodeling events and cell interactions. While TIMPs are a family of 4 secreted proteins that share overlapping substrate specificities of MMPs, RECK is unique in that it is a membrane-anchored MMP inhibitor that is embryonic lethal in mice. I used Xenopus laevis as a model organism to investigate the role of RECK as …

Effect Of N-Acetyl-L-Cysteine Prevention Or Intervention On Diet Induced Beta Cell Compensation And Dysfunction, Madison Wallace

Electronic Thesis and Dissertation Repository

Type 2 diabetes mellitus (T2DM) progression increases oxidative stress which contributes to beta cell compensation and eventual dysfunction. To investigate the role of antioxidant N-acetyl-L-cysteine (NAC) on beta cell function and pancreatic stellate cell activation (aSMA⁺) during early and late stages of compensation, NAC was used for preventative (p) and intervention (i) treatments in C57BL/6N mice fed a 60% kcal high-fat diet (HFD) for 8 or 22 weeks. Significantly improved glucose tolerance was observed at 22 weeks following pNAC treatment in HFD mice. Although 22-week HFD mice displayed hyperinsulinemia, beta cell hypertrophy, decreased beta cell PDX-1 nuclear localization, …

Formation Of A Vascular Regenerative Microenvironment Within Implantable Human Decellularized Adipose Tissue Bioscaffolds, Christopher Leclerc

Electronic Thesis and Dissertation Repository

Cellular therapies targeted at stimulating therapeutic angiogenesis in individuals with critical limb ischemia (CLI) have been under intense investigation. Hematopoietic progenitor cells (HPC) derived from umbilical cord blood have been previously shown to support limb revascularization in animal models of CLI, despite limited cell survival at the site of ischemia. This study attempted to improve HPC survival after transplantation and prolong pro-angiogenic function using human decellularized adipose tissue (hDAT) as a novel cell delivery platform. Compared to HPC conventionally grown on tissue-cultured plastic, hDAT scaffolds were shown to promote viability and proliferation of seeded HPC, and had cell- instructive effects …

The Role Of Tra1 In Cellular Stress Responses In Yeast: Implications For Human Diseases, Yuwei Jiang

Electronic Thesis and Dissertation Repository

Regulation of gene expression under stress conditions involves chromatin remodeling through post-translational modification of histones. One of these modifications, acetylation of lysine residues, regulates transcription initiation and is linked to a variety of essential cellular processes including cell cycle control, DNA repair, and importantly, activation of cellular stress response pathways. Dysregulation of histone acetylation has been observed in many stress-related diseases such as inflammatory diseases, cancer, neurodegenerative disorders, and fungal infections. Tra1 is the only essential component of both the highly conserved SAGA and NuA4 histone acetyltransferase (HAT) complexes that are responsible for acetylation of histones and other proteins. Tra1 …

Integrin-Linked Kinase Modulation Of Melanocytic Lineage Cells, Melissa Crawford

Electronic Thesis and Dissertation Repository

Melanocytes are specialized melanin-producing cells found in the skin, inner ear and heart. Melanocyte abnormalities cause many human disorders, including pigmentation defects, deafness, and melanoma. A better understanding of melanocyte biology is essential to address those disorders. Integrin-linked kinase (ILK) is a ubiquitous scaffold protein, essential for epidermal development. Importantly, the mechanisms by which ILK modulates the development and functions of melanocytic lineage cells remains unknown. To help address this void, I have developed cell-based models, as well as a reporter mouse model that allows tamoxifen-inducible Ilk gene inactivation specifically in melanocytic cells.

I observed that inactivation of Ilk in …

Hei-Oc1 Cochlear Cells As An In Vitro Model To Study The Role Of Connexins In Ototoxicity And Hearing Loss, Rianne Beach

Electronic Thesis and Dissertation Repository

Connexin 26 (Cx26) and Cx30 mediate the intercellular exchange of metabolites and ions within the cochlea in a process known as gap junctional intercellular communication (GJIC). Cochlear cell death and subsequent hearing loss can arise after treatment with ototoxic therapeutics and Cx26 mutant expression. We investigated the role of connexins and GJIC in the development of ototoxicity in HEI-OC1 cochlear-derived cells. The susceptibility of HEI-OC1 cells to aminoglycoside antibiotics and cisplatin-induced cell death was not influenced by the ablation of connexins and GJIC. However, the expression of mitochondrial apoptosis or ER stress markers was altered by the degree of GJIC. …

A Role For Shh And Bmp4 In Regulating The Dorsal-Ventral Patterning Of The Developing Pharyngeal Region, Alex Szpak

Electronic Thesis and Dissertation Repository

The pharynx is crucial to the survival of all vertebrates since it facilitates respiration by connecting the nasal and oral cavity to the larynx and digestion by connecting the oral cavity to the esophagus. The developing pharyngeal region displays dorsoventral patterning, and currently there is little information identifying the underlying mechanisms that regulate this patterning. This is in part due to the complexity of the developing pharyngeal region that requires contributions from all three germ layers along with neural crest cells. The expression profiles of Sonic Hedgehog (Shh) and Bone Morphogenetic Protein 4 (Bmp4) adjacent to the developing pharyngeal region …

Pregestational Diabetes Induced Congenital Heart Defects And Coronary Artery Malformations; Mechanisms And Preventative Therapies, Anish Engineer

Electronic Thesis and Dissertation Repository

Congenital heart defects (CHDs) arise from perturbations in complex molecular and cellular processes underlying normal embryonic heart development. CHDs are the most common congenital malformation, occurring in 1 to 5% of live births, and are the leading cause of pediatric mortality. Adverse genetic and environmental factors can impede normal cardiogenesis and increase the likelihood of CHDs. Pregestational maternal diabetes increases the risk of CHDs in children by more than four-fold. As the prevalence of diabetes rapidly rises among women of childbearing age, there is a need to investigate the mechanisms and potential preventative strategies for these defects. The aim of …

Maternal Nicotine Exposure Induces Congenital Heart Defects In The Offspring Of Mice, Elizabeth Greco

Electronic Thesis and Dissertation Repository

Congenital heart defects are the most prevalent birth defect, and maternal cigarette smoking is a known risk factor. Nicotine replacement therapies are recommended to pregnant women who smoke to aid in smoking cessation, as this alternative is thought to be much safer compared to cigarette smoking. However, these products contain nicotine, and the safety of nicotine on the developing heart is not well known. In this thesis, a mouse model was used to test the hypothesis that maternal nicotine exposure (MNE) during pregnancy leads to congenital heart defects and coronary artery defects in the offspring of mice. MNE resulted in …

Absence Of Activating Transcription Factor 3 Reduces Severity Of Recurrent Pancreatitis And K-Ras Mediated Pdac In Mice, Nawab Azizi

Electronic Thesis and Dissertation Repository

Pancreatic ductal adenocarcinoma (PDAC) is a terminal cancer projected to become the leading cause of cancer-related deaths in North American by the year 2030. Constitutive activation of KRAS is seen in >90% of PDAC cases. In addition to oncogenic KRAS activity, pancreatic injury is key contributor to PDAC initiation and progression. Activating transcription factor 3 (ATF3) is required for the formation of pre-neoplastic lesions in acute pancreatitis. However, unlike recurrent or chronic forms of pancreatitis, acute pancreatitis is not predictive of PDAC. Therefore, the goal of this thesis is to determine the role of ATF3 in recurrent pancreatitis and PDAC. …

Elucidating The Mechanism Of Nuak1 Function In Ovarian Cancer Metastasis, Jamie Fritz

Electronic Thesis and Dissertation Repository

Epithelial ovarian cancer (EOC) has a unique mode of metastasis, where upon shedding from the primary tumor, cells form spheroids and spread through the peritoneal cavity. There is a need to elucidate pathways driving spheroid formation to identify novel therapeutic targets. We previously showed that LKB1 is required for EOC metastasis. Using multiplex inhibitor bead-mass spectrometry, we identified NUAK1 as a top LKB1 substrate candidate. We confirmed LKB1 maintains NUAK1 phosphorylation and expression. NUAK1KO cells had lower cell adhesion and generated spheroids with reduced integrity. We identified a cell attachment pathway that was enriched in parental compared with NUAK1 …

Ampk Signalling As A Regulator Of Autophagy In A Model Of Ovarian Tumour Dormancy, Jeremi Laski

Electronic Thesis and Dissertation Repository

One of the hallmarks of epithelial ovarian cancer (EOC) metastasis lies in the process of spheroid formation, whereby tumour cells aggregate into 3D structures. Previous literature suggests that as EOC cells form spheroids they undergo bioenergetic stress, activate AMP-activated protein kinase (AMPK) signaling, and thereby force cells to enter a metabolically quiescent state. We have previously shown that EOC spheroids up- regulate autophagy, a process that provides energy during starvation conditions. Herein, I examined the role of AMPK-mediated signaling regulation of autophagy in a model of ovarian tumour dormancy. Attenuation of AMPK signaling in EOC spheroids resulted in reduction of …

Oleic Acid Attenuates Palmitic Acid-Induced Impairments In Mouse Blastocyst Development, Maisoon Dafalla Yousif

Electronic Thesis and Dissertation Repository

Palmitic acid (PA) and oleic acid (OA) are the most abundant saturated and monounsaturated non-esterified fatty acids, respectively, in serum. The consequences of PA and OA treatment on preimplantation mouse embryo development were investigated. It was hypothesized that OA treatment from the two-cell stage onwards would attenuate PA-induced reductions in mouse blastocyst development. PA significantly reduced development to the blastocyst stage. Transcript analysis revealed that PA increased ER stress pathway markers activating transcription factor 3 and C/EBP homologous protein transcripts, and XBP1 splicing. Co-culture of PA-treated embryos with OA prevented this PA-induced ER stress and rescued blastocyst development. Cytoplasmic lipid …

Phenotypic And Metabolic Plasticity In Canine Cellular Reprogramming, Ian C. Tobias

Electronic Thesis and Dissertation Repository

Embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) derived from large mammals reproduce few characteristics displayed by rodent or human counterparts. This is complicated by the inherent plasticity of mammalian ESC/iPSC cultures that resemble a variety of developmental stages including the naïve and primed pluripotent states. Defining the extrinsic signals that modulate the developmental identity of canine ESC/iPSC (i.e. primed versus naïve) will improve knowledge integration with more sophisticated rodent and primate research. In this thesis, I sought to determine if manipulation of the culture environment can promote nuclear and metabolic reprogramming of canine cell lines towards a …

Hyperactive Torc1 Sensitizes Yeast Cells To Endoplasmic Reticulum Stress By Compromising Cell Wall Integrity, Khadija Ahmed

Electronic Thesis and Dissertation Repository

The disruption of protein folding homeostasis in the endoplasmic reticulum (ER) results in an accumulation of toxic misfolded proteins and activates a network of signaling events collectively known as the unfolded protein response (UPR). While UPR activation upon ER stress is well characterized, how other signaling pathways integrate into the ER proteostasis network is unclear. Here, I sought to investigate how the target of rapamycin complex 1 (TORC1) signaling cascade acts in parallel with the UPR to regulate ER stress sensitivity. Using S. cerevisiae, I found that TORC1 signaling is attenuated during ER stress and constitutive activation of TORC1 …

Cell Recruitment And Differentiation In Wound Repair, John Walker

Electronic Thesis and Dissertation Repository

Chronic wounds represent a major health problem primarily affecting patients with underlying pathologies including diabetes and vasculopathy. These wounds are characterized by an impaired transition from the inflammatory to the proliferative phase of healing. In this thesis, the complex relationship between cells and their microenvironment was explored in the context of wound healing by investigating how intrinsic differences in fibroblasts affect their response to external stimuli, and how extracellular matrix (ECM) proteins modify cellular responses during acute wound repair. First, it was hypothesized that different subpopulations of fibroblasts could be identified in cutaneous tissue based on divergent embryonic expression patterns …

Alpha-Synuclein Toxicity Is Caused By Mitochondrial Dysfunction, Michael G. Tauro

Electronic Thesis and Dissertation Repository

Parkinson’s disease (PD) is the second most common neurodegenerative disorder, affecting roughly 1% of the population over the age of sixty years. Alpha-synuclein (aSyn) is a protein implicated in both familial and idiopathic forms of PD, yet despite the wealth of data implicating aSyn as a causative agent in PD, the mechanisms underlying its toxicity remain mostly unknown. Mitochondrial dysfunction is a major hallmark of PD, yet there is only limited evidence linking aSyn toxicity to mitochondrial dysfunction. My study establishes a novel aSyn model in respiring yeast cells, which allows me to explore how aSyn affects mitochondrial homeostasis and …