Tumour suppressor 73 (Trp73) gene shares structural and functional homology with p53. Trp73 encodes multiple isoforms with opposing effects. The TAp73 isoform is a transcription factor and is classified as a tumour suppressor where as the DNp73 isoform is a putative oncogene. Imbalance of the two opposing isoforms has been reported in human malignancies including ovarian cancer. TAp73 deficiency results in a dramatic increase in the DNp73 isoform, mimicking the altered isoform balance observed in human ovarian cancer. The effects of TAp73 deficiency in mouse OSE cells were assessed. TAp73 deficiency is not sufficient to induce ovarian cancer in mice. However, TAp73 deficiency compromises cellular proliferation in OSE cell lines. Furthermore, expression of p73 isoforms and epithelial markers is altered in TAp73 deficient cell line. Further studies are needed to determine if TAp73 deficiency in conjunction with other molecular alterations can mediate transformation of OSE cells leading to ovarian cancer.

Type 2 diabetes is characterized by insulin resistance and pancreatic β-cell failure. Insulin resistance leads to increased insulin demand, which can lead to increased proinsulin misfolding in the endoplasmic reticulum (ER). The accumulation of the misfolded proteins in the ER can cause ER stress, which can lead to pancreatic β-cell dysfunction. Cells respond to ER stress by the unfolded protein response (UPR), which increases protein folding capacity and causes degradation of misfolded proteins. Using a pancreatic β-cell model of induced misfolded proinsulin expression (proinsulin-C96Y tagged with GFP) we discovered that one of the most highly induced genes was stromal cell-derived factor 2 like 1 (SDF2L1). SDF2L1 is an ER localized soluble protein with an as yet unknown function. In this thesis I examined the potential role of SDF2L1 in pancreatic β-cells in ER stress conditions.

Exposure to hyperoxia has been shown to have detrimental effects on newborn hemodynamics and antioxidant activities in males. These effects are unknown in the newborn female, despite clinical data showing that in children undergoing surgery females have poorer outcomes. This thesis explored whether newborn females respond worse than males to hyperoxia, specifically in hemodynamics and antioxidant activity. Hemodynamic results showed continual decreases in DBP and MAP, increases in HR, decreases in SBP, and increases in PP, which were experienced earlier and to a greater degree in the female. Additionally, reduced antioxidant activity seen in newborns was worse in females in both the heart and skeletal muscle. These results suggest that in response to hyperoxia, newborn females are at a hemodynamic disadvantage when compared to males, which may be a contributing factor for why female children are at a higher risk during surgery or medical management involving hyperoxia.

Short-chain fatty acids (SCFAs) are anaerobic bacterial metabolites. Cystic fibrosis (CF) lung disease is a condition caused by mutations in the cystic fibrosis transmembrane conductange regulator (CFTR) gene and is characterized by persistent lung inflammation and bacterial colonization. We measured the concentrations of SCFAs in sputum of patients with CF and tested the effect of these compounds on bacterial growth. Furthermore we found that SCFAs can influence the inflammatory protein expression and cytokine release in airway epithelial cells. SCFAs differentially alter cytokine release in CF bronchial epithelial cells (CFBE) compared to CFBE expressing wild-type CFTR. We also studied the effect of SCFAs in an acute lung injury model in BALB/cJ mice and found that intratracheally administered SCFAs can affect the inflammatory environment of the airways in vivo. We conclude that SCFAs may be important in the airways and that further investigation is warranted to understand their effects on inflammation and infection.

Blood CORT is commonly used to assess stress in rodents, but sampling can trigger a rapid stress response. This study aims to identify whether faecal CORT metabolites (FCM) can reflect changes in CORT induced by 7-day food restriction (FR) and an ACTH challenge. Blood and 24hr faecal samples were collected at baseline and Day 7 for control (n=8) and FR (n=10) rats. On Day 8, after a baseline blood sample, an ACTH injection was administered and followed by blood and fecal sampling. Results showed increased serum CORT and FCM in response to FR. Increased adrenal sensitivity with FR was illustrated by a greater increase in serum CORT compared to control in response to ACTH. Lastly, although it appeared that ACTH induced an increase in FCM in FR and control, only the latter reached statistical significance. Thus FCM might be better suited for quantifying chronic rather than acute changes in CORT.

Gonadotropin-Releasing Hormone (GnRH), the master regulator of the reproductive axis, is hypothesized to play an essential role in directly sensing changes in energy levels to maintain fertility. There are a number of GnRH cell lines that have been utilized to study reproductive function; however these have been embryonic in origin and clonally derived. To investigate the cellular mechanisms underlying glucose responsiveness in GnRH neurons, we generated a novel GnRH cell line through immortalization, and fluorescent activated cell (FAC)-sorting of hypothalamic primary culture taken from a GnRH-GFP transgenic mouse. The mHypoA-GnRH/GFP neurons express a complement of markers of fully differentiated GnRH neurons. Glucose induces neuronal activation of mHypoA-GnRH/GFP neurons and glucose metabolism initiates an AMP-Protein Kinase (AMPK)-dependent mechanism to exert transcriptional and secretory control of GnRH. These findings support the use of this novel GnRH cell model for defining components involved in cellular and molecular action of glucose in GnRH neurons.

Skin-derived precursors (SKPs) are multipotent dermal stem cells that reside within a hair follicle niche and that share properties with embryonic neural crest precursors. Here, we have asked whether SKPs and their endogenous dermal precursors originate from the neural crest or whether, like the dermis itself, they originate from multiple developmental origins. To do this, we used two different mouse Cre lines that allow us to perform lineage tracing: Wnt1-cre, which targets cells deriving from the neural crest, and Myf5-cre, which targets cells of a somite origin. By crossing these Cre lines to reporter mice, we show that the endogenous follicle-associated dermal precursors in the face derive from the neural crest, and those in the dorsal trunk derive from the somites, as do the SKPs they generate. Despite these different developmental origins, SKPs from these two locations are functionally similar, even with regard to their ability to differentiate into Schwann cells, a cell type only thought to be generated from the neural crest. Analysis of global gene expression using microarrays confirmed that facial and dorsal SKPs exhibit a very high degree of similarity, and that they are also very similar to SKPs derived from ventral dermis, which has a lateral plate origin. However, these developmentally distinct SKPs also retain differential expression of a small number of genes that reflect their developmental origins. Thus, an adult neural crest-like dermal precursor can be generated from a non-neural crest origin, a finding with broad implications for the many neuroendocrine cells in the body.

INTRODUCTION: Platelets are involved in the inflammatory and thrombotic complications associated with atherosclerosis. Human neutrophil peptides (HNP), released from activated neutrophils, demonstrate inflammatory effects related to lesion development. HNP bind the low-density lipoprotein receptor (LR) family member LRP1 and LRP8 is the only member on platelets. HYPOTHESIS: HNP enhance platelet activation and aggregation through interactions with LRP8. METHODS: Platelet activation and aggregation in response to HNP were determined using flow cytometry and aggregometry. Activation was also examined in the presence of recombinant LRP8 and in LRP8 knockout platelets. RESULTS: HNP activate platelets as determined by P-selectin expression and the formation of microparticles. HNP sensitize platelets enhancing their aggregatory response to ADP. Lastly, LRP8 plays a role in HNP-induced platelet activation. CONCLUSIONS: With an improved understanding of the mechanism by which HNP induce platelet activation, we may be able to devise therapeutic strategies to treat patients with cardiovascular diseases.

Skin-derived precursors (SKPs) are multipotent dermal stem cells that reside within a hair follicle niche and that share properties with embryonic neural crest precursors. Here, we have asked whether SKPs and their endogenous dermal precursors originate from the neural crest or whether, like the dermis itself, they originate from multiple developmental origins. To do this, we used two different mouse Cre lines that allow us to perform lineage tracing: Wnt1-cre, which targets cells deriving from the neural crest, and Myf5-cre, which targets cells of a somite origin. By crossing these Cre lines to reporter mice, we show that the endogenous follicle-associated dermal precursors in the face derive from the neural crest, and those in the dorsal trunk derive from the somites, as do the SKPs they generate. Despite these different developmental origins, SKPs from these two locations are functionally similar, even with regard to their ability to differentiate into Schwann cells, a cell type only thought to be generated from the neural crest. Analysis of global gene expression using microarrays confirmed that facial and dorsal SKPs exhibit a very high degree of similarity, and that they are also very similar to SKPs derived from ventral dermis, which has a lateral plate origin. However, these developmentally distinct SKPs also retain differential expression of a small number of genes that reflect their developmental origins. Thus, an adult neural crest-like dermal precursor can be generated from a non-neural crest origin, a finding with broad implications for the many neuroendocrine cells in the body.

Atrial fibrillation (AF) is the most common supraventricular tachyarrhythmia that can present without cardiovascular disease (lone AF). Frequent high-intensity endurance exercise is a risk factor for lone AF, and the pathophysiology of AF induced by intense endurance exercise is unknown. We found that after 6 weeks of intense swimming and running, mice were far more susceptible to AF, but not ventricular arrhythmias. Exercise induced atrial fibrosis, inflammation and slowed conduction without detectible changes in ventricles. Since AF is associated with stretch and since a tumor necrosis factor-α (TNFα) is a mechanosensitive inflammatory factor, mice were treated with the TNFα inhibitor etanercept. Etanercept treatment blocked inflammation, fibrosis, and AF vulnerability in the exercised mice. Consistent with these findings, we found that exercise caused large elevations in atrial pressures. Our findings support the conclusion that mechanical loading of atria during exercise induces TNFα release, leading to structural remodeling and enhanced AF vulnerability.