http://hdl.handle.net/1974/848 2013-05-19T17:29:45Z 2013-05-19T17:29:45Z Brisson, DEVIN http://hdl.handle.net/1974/7578 2012-10-05T05:05:15Z 2012-10-04T04:00:00Z

Title: Higher brain neurons succumb to acute stroke-like insult while lower brain neurons strongly resist Authors: Brisson, DEVIN Abstract: Pyramidal neurons (PyNs) in ‘higher’ brain are highly susceptible to acute stroke injury yet ‘lower’ brain regions better survive global ischemia, presumably because of better residual blood flow. Here we show that projection neurons in ‘lower’ brain regions of hypothalamus and brainstem intrinsically resist acute stroke-like injury independent of blood flow in the brain slice. In contrast `higher` projection neurons in neocortex, hippocampus, striatum and thalamus are highly susceptible. In live brain slices from rat deprived of oxygen and glucose (OGD), we imaged anoxic depolarization (AD) as it propagates through these regions. AD, the initial electrophysiological event of stroke, is a depolarizing front that drains residual energy in compromised gray matter. The extent of AD reliably determines ensuing damage in higher brain, but using whole-cell recordings we found that all CNS neurons do not generate a robust AD. Higher neurons generate strong AD and show no functional recovery in contrast to neurons in hypothalamus and brainstem that generate a weak and gradual AD. Most dramatically, lower neurons recover their membrane potential, input resistance and spike amplitude when oxygen and glucose is restored, while higher neurons do not. Following OGD, new recordings could be acquired in all lower (but not higher) brain regions, with some neurons even withstanding multiple OGD exposure. Two-photon laser scanning microscopy confirmed neuroprotection in lower, but not higher gray matter. Specifically pyramidal neurons swell and lose their dendritic spines post-OGD, whereas neurons in hypothalamus and brainstem display no such injury. Exposure to the Na+/K+ ATPase inhibitor ouabain (100 μM), induces depolarization similar to OGD in all cell types tested. Moreover, elevated [K+]o evokes spreading depression (SD), a milder version of AD, in higher brain but not hypothalamus or brainstem so weak AD correlates with the inability to generate SD. In summary, overriding the Na+/K+ pump using OGD, ouabain or elevated [K+]o evokes steep and robust depolarization of higher gray matter. We show that this important regional difference can be largely accounted for by the intrinsic properties of the resident neurons and that Na+/K+ ATPase pump efficiency is a major determining factor generating strong or weak spreading depolarizations. Description: Thesis (Ph.D, Anatomy & Cell Biology) -- Queen's University, 2012-10-02 17:59:20.589

2012-10-04T04:00:00Z ARMSTRONG, DAVID http://hdl.handle.net/1974/7559 2012-10-02T05:02:55Z 2012-10-01T04:00:00Z

Title: DEVELOPMENTAL ORIGINS OF CARDIOVASCULAR DISEASE: ATRIAL NATRIURETIC PEPTIDE GENE DISRUPTED MICE AS A MODEL OF GESTATIONAL HYPERTENSION Authors: ARMSTRONG, DAVID Abstract: Introduction: Developmental origins of disease refers to the theory that adverse maternal environments influence fetal development and the risk of cardiovascular disease (CVD) in adulthood. To test the hypothesis that gestational hypertension influences the development of CVD in offspring, a novel experimental paradigm was developed using atrial natriuretic peptide gene disrupted mice (ANP-/-). The objective of this thesis was to determine the effect of gestational hypertension on cardio-renal function in offspring. Methods: ANP+/+ females were crossed with ANP-/- males (yielding ANP+/-WT offspring) and ANP-/- females with ANP+/+ males (yielding ANP+/-KO offspring). Previous work has established that ANP-/- dams are hypertensive during pregnancy. Offspring gene expression was measured using qPCR. Offspring arterial blood pressure (BP) was measured with a non-invasive tail cuff system. Offspring left ventricular (LV) function was examined using echocardiography (ECHO). Offspring were treated with normal salt (NS) or high salt (HS) chow for five weeks to assess salt-sensitivity. Daily injections of isoproterenol (ISO) were used to induce cardiac stress in offspring. Collagen deposition was assessed using Masson’s trichrome and picrosirius red staining. Results: Absence of maternal ANP had no effect on either litter size or offspring growth, but caused significant LV hypertrophy in offspring, with no change in LV function. Treatment with ISO resulted in myocardial fibrosis and significant LV diastolic dysfunction with a restrictive filling pattern (increased E/A ratio and E/e’) only in ANP+/-KO offspring. Furthermore, absence of maternal ANP was associated with salt-resistant BP in offspring. Conclusions: Gestational hypertension using the ANP-/- mouse model results in a salt-resistant phenotype in offspring, as well as significant cardiac hypertrophy and an adverse response to activation of the sympathetic nervous system in adult offspring. These data suggest that adverse maternal environments may increase the risk of cardiovascular disease in offspring later in life. Description: Thesis (Ph.D, Anatomy & Cell Biology) -- Queen's University, 2012-09-18 16:12:01.147

2012-10-01T04:00:00Z Smallwood, Chelsea http://hdl.handle.net/1974/7374 2012-08-15T22:54:04Z 2012-08-15T04:00:00Z

Title: THE EFFECT OF HYPOXIA INDUCIBLE FACTOR-1 ON THE EXPRESSION OF THE COINHIBITORY LIGANDS B7-H3 AND B7-H1 IN CANCER: RELEVANCE TO CANCER IMMUNE ESCAPE Authors: Smallwood, Chelsea Abstract: The interactions between tumour cells and cells of the immune system are important in the natural evolution of cancer, and the acquired immune system plays an integral role in cancer immune escape. B7-H3 and B7-H1 ligands provide coinhibitory signals to T cells resulting in T cell anergy or apoptosis and their expression has been shown to increase in cancer cells. Tumour hypoxia (oxygen concentration below physiological level) is a major contributor to the spread of cancer and resistance to radiation and chemotherapy. We proposed that hypoxia results in the upregulation of the B7 molecules B7-H3 and B7-H1. Furthermore, studies in our laboratory have shown that acquisition of malignant properties in tumour cells exposed to hypoxia can be inhibited by low concentrations of nitric oxide mimetic agents such as glyceryl trinitrate (GTN). Using cultured breast and prostate cancer cells, we investigated whether the hypoxia-inducible factor HIF-1α, would mediate an upregulation of these ligands. Using a mouse model, we investigated the effect of GTN on tumour growth in vivo. For the in vitro studies, we exposed MDA-MB-231 and MCF-7 breast cancer cells and DU-145 prostate cancer cells to standard culture conditions, hypoxic conditions, or 100 μM CoCl2 (stabilizes HIF-1α) for 24 hours. Our findings indicate that B7-H3 mRNA was upregulated in hypoxia (P = 0.0101). Contrary to our hypothesis, B7-H3 protein was not upregulated in hypoxia. Interestingly, increased B7-H1 protein expression correlated with increased HIF-1α expression (r2=0.48, P<0.0001), and HIF-1α bound to the hypoxia response element (HRE) of B7-H1. These results indicate a role for HIF-1α in the upregulation of B7-H1 levels in MDA-MB-231 cells. While in vitro studies indicated no effect of GTN, a study using female BALB/c mice injected with 4T1 mammary carcinoma cells resulted in a decrease in tumour volume in the GTN treated mice. Together, these results indicate a novel role for HIF-1α in the up-regulation of B7-H1 on cancer cells, thus potentially contributing to immune escape of cancer cells and additionally, a role for GTN as a possible breast cancer therapy. Description: Thesis (Master, Anatomy & Cell Biology) -- Queen's University, 2012-08-15 11:10:06.237

2012-08-15T04:00:00Z VIRANI, SOPHIA http://hdl.handle.net/1974/6920 2011-12-23T05:58:48Z 2011-12-22T05:00:00Z

Title: Role of Stromal Cell-Derived Factor-1 in Neoangiogenesis in Endometriosis Lesions Authors: VIRANI, SOPHIA Abstract: Endometriosis affects 5-10% of women and is characterized by the growth of endometrial tissue outside of the uterus. Treatment for endometriosis primarily focuses on symptom relief, is short term with severe side effects and often leads to recurrence of the condition. Establishing new blood supply is a fundamental requirement for endometriosis lesions growth. This has led to the idea that antiangiogenic therapy may be a successful approach for inhibiting endometriosis. Recent evidence indicates that endothelial progenitor cells (EPCs) contribute to neoangiogenesis of endometriotic lesions. These EPCs are recruited to the lesion site by stromal cell-derived factor-1 (SDF-1). We hypothesize that SDF-1 is central to the neoangiogenesis and survival of endometriotic lesions and that administration of SDF-1 blocking antibody will inhibit lesion growth by inhibiting angiogenesis in a murine model of endometriosis. Immunohistochemistry for SDF-1 and CD34 was performed on human endometriosis and normal endometrial samples. Quantification of SDF-1 and EPCs was performed in the blood of endometriosis patients and controls using ELISA and flow cytometry, respectively. A new mouse model of endometriosis was developed using BALB/c-Rag2-/-/IL2rg-/- mice to investigate role of SDF-1 in neoangiogenesis. Either SDF-1 blocking antibody or an isotype control was administered on a weekly basis for four weeks. Weekly samples of peripheral blood from mice were analyzed for SDF-1, other cytokines of interest and EPCs. Mice were euthanized at seven weeks to observe lesion growth and blood vessel development. Our results indicate overabundance of SDF-1 and CD34+ progenitor cells in human endometriotic lesions compared to eutopic endometrium. In the mouse model, SDF-1 and circulating EPC levels decreased from pre-treatment levels after one week, and remained constant over the course of the treatment in both SDF-1 blocking antibody and isotype control groups. In the SDF-1 blocking group, reduced vascularity of lesions, identified by immunofluorescence staining for CD31, was revealed compared to isotype controls. These findings suggest that SDF-1 may be responsible for CD34+ progenitor cell recruitment to the neoangiogenic sites in endometriosis. Blocking of SDF-1 reduces neovascularization of human endometriotic lesions in a mouse model. Further studies on blocking SDF-1 in combination with other antiangiogenic agents are needed. Description: Thesis (Master, Anatomy & Cell Biology) -- Queen's University, 2011-12-21 19:34:43.054

2011-12-22T05:00:00Z