<p>Globalization scholars have disagreed about the effects of globalization on the production and reproduction of difference: Do fundamental differences endure, do cultures converge, or is there hybridization? This dissertation analyzes the durability of distinct medical cultures in two technologically advanced healthcare systems that rely on an evidence-based, biomedical approach. Durability refers to the tendency to maintain or develop diverse, even idiosyncratic, practices and beliefs--even as the forces of globalization are perceived to be pressing health practices everywhere toward a single global standard. To do so, this dissertation offers a comparative, empirically based argument using the case of type 2 diabetes in the U.S. and Japan. As an inductive, theory-constructing project, the argument has at its foundation 11 months of ethnographic field work in Japanese hospitals and clinic exam rooms, 115 semi-structured interviews with patients and biomedical health practitioners in Japan, and 25 interviews with American health care providers and patients. I argue that physicians in both research sites, Okayama, Japan and North Carolina, USA, practice empirical biomedicine, but that empirical biomedicine is not all there is to biomedical practice. Practicing physicians in both contexts act not only on increasingly globalized professional standards, but also on local knowledge, on their own explanatory models for type 2 diabetes, and in reaction to local patient populations' explanatory models. Further, local knowledge and patient interactions shape the ways in which practicing physicians interpret global standards and best practices. Occasionally, they may even be reshaped beyond recognition without interfering with physicians' self-evaluation as participants in a universal, standardized scientific project. The interaction of globalizing standards of practice, local knowledge, and local explanatory models of illness can result in dramatically divergent medical practice across different social contexts--in this case, the U.S. and Japan.</p> / Dissertation

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Glucose transporter GLUT4 allows glucose uptake into muscle and adipose cells. Insulin promotes recruitment and plasma membrane insertion of GLUT4 vesicles that can recycle constitutively. Obesity and type 2 diabetes mellitus are associated with defects in insulin-induced GLUT4 recruitment. Knowledge of alternative modes and steps of GLUT4 traffic in L6-GLUT4myc muscle cells may reveal possible targets for therapeutic intervention in insulin-resistant patients. Hypertonicity and Platelet Derived Growth Factor also increase surface GLUT4 levels but it was unknown if they tap on the same intracellular GLUT4 depots as insulin. We explored whether GLUT4 vesicles recycle using different compartments and mechanisms for the surface gain elicited by each stimulus. We hypothesized that all vesicle fusion steps require NSF but depend on individual v-SNAREs. Specifically, we tested effects of ATPase-deficient NSF or VAMP7 siRNA transfections, and endosomal ablation on GLUT4 traffic. We show that VAMP7 was required for basal and hypertonic recycling, while VAMP2 is exclusively used in response to insulin. As insulin action bifurcates downstream of phosphatidylinositol 3-kinase, we also hypothesized that the Rac-to-actin and Akt-to-AS160 branches regulate distinct GLUT4 traffic steps. For this we determined GLUT4myc localization in rounded myoblasts relative to a surface marker. Interfering with Rac, actin dynamics or actin-binding α-actinin4 maintained GLUT4 in a perinuclear region even under insulin-stimulation. Interfering with AS160 allowed significant GLUT4 accumulation beneath the membrane, but not fusion. We propose that actin dynamics and α-actinin4 are required for cortical GLUT4 tethering mechanisms, and AS160 contributes to GLUT4 docking/fusion. We confirmed that VAMP2 facilitates GLUT4 fusion, as tetanus toxin-based cleavage did not inhibit peripheral GLUT4 recruitment. Finally, AS160 targets Rab8A and Rab14 in muscle respectively affected GLUT4 availability for membrane fusion, and basal GLUT4 retention. This work will lead to future testing of strategies to selectively enhance vesicle availability, tethering, or surface fusion, for bypassing insulin resistance.

During atherosclerosis and restenosis, there is an extensive amount of collagen synthesis and degradation. Changes in the types of collagen present can have profound effects on vascular smooth muscle cell (SMC) proliferation and migration. Type VIII collagen, which is normally present at low levels within the mature vascular system, is greatly increased during atherogenesis. The central theme of this thesis is to determine the role of type VIII collagen in the pathogenesis of atherosclerosis and restenosis. In the first study, we demonstrated the importance of type VIII collagen in SMC migration and proliferation. SMCs from type VIII collagen-deficient mice display increased adhesion and decreased spreading, migration, and proliferation compared to SMCs from wild-type mice. Treatment of SMCs from type VIII collagen-deficient mice with exogenous type VIII collagen can rescue the defects. In the second study, we determined that type VIII collagen exerts its effects through regulation of MMP-2 expression. Type VIII collagen-deficient SMCs have decreased levels of MMP-2 and are impaired in chemotaxis toward PDGF-BB and in their ability to contract thick collagen gels. We found that decreasing endogenous MMP-2 levels in normal SMCs or adding exogenous collagen to type VIII collagen-deficient SMCs is sufficient to recapitulate the type VIII collagen-deficient or wild-type SMC phenotype, respectively. In the third study, we investigated the contribution of type VIII collagen to intimal hyperplasia after mechanical injury in the mouse. We found that type VIII collagen-deficient mice display a 35% reduction in intimal hyperplasia and attenuated vessel remodeling after femoral artery wire injury, establishing a role for type VIII collagen in restenosis. The results of the work presented in this thesis demonstrate that production of type VIII collagen confers an SMC phenotype with a greater propencity for proliferation and migration. These effects are in part mediated through regulation of MMP-2 expression and activation. We conclude that the increases in type VIII collagen production that occur during atherosclerosis and restenosis contribute to the capacity of SMCs to alter the existing extracellular matrix in a manner which permits enhanced migration.

Type II diabetes and hyperinsulinemia are associated with increased risks of developing colorectal cancer (CRC). Detailed mechanisms underlying this correlation, however, are yet to be explored. The present study demonstrates that insulin increases the expression of proto-oncogenes c-Myc and cyclin D1 via both translational and transcriptional mechanisms. We show here that insulin stimulates c-Myc gene translation via an Akt/PKB-dependent mechanism involving the mTOR signaling pathway. More importantly, we show for the first time that transcriptional stimulation of c-Myc and cyclin D1 expression by insulin involves a novel Akt/PKB-independent signal crosstalk between insulin and canonical Wnt signaling pathways. We then identified p21-activated protein kianse 1 (PAK-1) as a novel mediator for insulin and Wnt/beta-catenin (b-cat) molecular crosstalk, involving MEK/ERK signaling. Furthermore, we found that insulin treatment leads to increased b-cat phosphorylation at Ser675, and this is associated with increased b-cat nuclear content and increased b-cat interaction with Tcf/Lef-binding elements (TBEs) of the human c-Myc gene promoter. Lastly, we demonstrated that insulin signaling directly alters the expression levels of components of the Wnt signaling pathway, including fizzled homology 4 (Fdz-4) and TCF7L2 (=TCF-4). This study not only demonstrated the existence of signaling crosstalk between insulin and canonical Wnt signaling pathways at multiple levels, it reveals molecular mechanisms for observed correlation between CRC and hyperinsulinemia. The growing evidence implicating PAK-1 in various human tumorigenesis has emerged PAK-1 as a potential therapeutic target. Our discovery of PAK-1 functioning as a novel central mediator for insulin and Wnt signaling crosstalk in intestinal cells suggests that PAK-1 may potentially be a good target candidate for treating patients with CRC, especially those who have Type II diabetes or experience hyperinsulinemia.

Vaccination has proven a cost-effective method of managing infectious diseases, but attempts to develop an effective fetal vaccine have proven difficult due to the immaturity of the immune system and the propensity of the developing immune system to induce tolerance to immunizing antigens. This thesis is concerned with the induction of specific immunity in the second trimester ovine fetus using the oral DNA immunization method. In utero oral delivery of naked DNA plasmid was selected as the method of immunization due to previous successes in the third trimester ovine fetus and the immunostimulatory properties of the bacterial DNA backbone, which may help overcome developmental tolerance. Transfection and expression studies in the third trimester ovine fetus revealed the oral mucosal epithelium as the primary site of transgene expression and functionally active antigen was also localized to lymph nodes draining the oral cavity. Efficient transfection and expression of plasmid following oral delivery was specific to the fetus and correlated with a lesser degree of epithelial differentiation. Oral DNA delivery in the second trimester resulted in detection of transgene activity in 100% of treated fetuses and the level of transgene activity was greater than in fetuses treated in the mid-third trimester. Using a plasmid encoding the gene for bovine herpesvirus-1 truncated glycoprotein D (tgD), immunization studies were then conducted in the second trimester fetus. A new lower age limit for fetal immunization was established at 55-60 days gestation (gestation period is 148 days), which coincides with the appearance of lymphocytes in peripheral tissues. Antigen-specific antibody, interferon-× responses and/or neonatal anamnestic responses were detected in 66% of fetuses immunized between 55 and 84 days gestation. The duration of fetal primary immune responses was equivalent to that achieved in young lambs following optimized DNA vaccination, but the magnitude of fetal immune responses was limited. The persistence of immune memory from the second trimester to birth was consistent with experimental data which showed that the duration of immune memory had a stronger correlation to the duration, as compared to the magnitude, of the primary antibody response. Overall, the experiments within showed that oral DNA immunization of the early second trimester fetus is feasible and not associated with the induction of tolerance. These findings suggest that it may be possible to protect against mother-to-child transmission of infectious pathogens by targeting protection at the level of the fetus.

The increasing input costs for traditional agriculture has led land owners and producers in search of alternative opportunities to increase on-farm income. Replacing agricultural crops with short rotation woody species such as hybrid poplar trees is a form of agroforestry. The objectives of this project were to evaluate: 1) a suitable planting stock for hybrid poplar, 2) the effect of nitrogen (N) fertilizer application and pruning on hybrid poplar growth and, 3) the response of four hybrid poplar clones to fertilizer application and their suitability in the boreal transition ecoregion of Saskatchewan. <p>Two trials were established near Meadow Lake, Saskatchewan where three stock types (cuttings, root cuttings and rooted plugs) of Walker poplar were planted into former alfalfa and pasture fields. Trees were pruned each spring to remove multiple leaders and fertilized in year 2 with 100 kg N ha-1. The presence of roots on rooted cutting and plug stock types was beneficial in terms of hybrid poplar growth and survival. Trees grown from planting stock without roots had survival rates between 32-37% whereas, the survival of trees with roots at the time of planting ranged from 62-81% after two years of growth. Trees that were planted as a rooted stock were 3.5 to 4.2 times greater in height and 4.0 to 5.6 times greater in root collar diameter than trees planted as an un-rooted stock type. The application of fertilizer N decreased tree volumes by 31% at the Alfalfa site and had no effect on tree growth at the Pasture site. The total amount of fertilizer N recovered by the hybrid poplar trees ranged from 1-3% at the Alfalfa site and 3-5% at the Pasture site. <p>The second study involved planting four clones of hybrid poplar (Hill, Katepwa, Walker and WP-69) at the same two sites and applying fertilizer at rates of 0, 150 and 300 kg N ha-1 the first two years. Following the second growing season, Katepwa and WP-69 clones had the highest tree volumes of 750 and 1147 cm3 of the four clones evaluated. The Walker clone had the poorest survival rates (52-56%) compared to the other three clones (> 90% survival). Foliar N levels were not correlated with tree height at the Alfalfa (p=0.1326) or the Pasture (p=0.1063) sites. The relationship between foliar P concentration and tree height was more pronounced during July at the Alfalfa site with an r2 value of 0.7102. The N:P ratios for foliar tissue decreased with increasing fertilizer N application during August at the Alfalfa site. Foliar N:P ratios were the same among fertilizer and clone treatments at the Pasture site in August. <p>Results from this study suggest that rooted stock types increase the successful establishment of hybrid poplar plantations. However, application of N fertilizer may not increase growth of trees if soil N is adequate. Other soil nutrients need to be measured prior to fertilization to determine what nutrients may be limiting plant growth.

Methylglyoxal (MG) is a reactive á-oxoaldehyde and a glucose metabolite. Previous studies in our laboratory have shown that MG induces the production of reactive oxygen species (ROS), such as superoxide (O2.-), nitric oxide (NO) and peroxynitrite (ONOO-), in vascular smooth muscle cells (VSMCs, A-10 cells). However, the effect of endogenous MG and mechanisms of MG-induced oxidative stress have not been thoroughly explored. The present study investigated fructose (a precursor of MG)- induced ONOO- formation in A-10 cells and whether this process was mediated via endogenous MG formation; roles of MG in regulating mitochondrial ROS (mtROS) production and mitochondrial functions in A-10 cells; and effect of MG on neutrophils in patients with type 2 diabetes mellitus (T2DM). Fructose induced intracellular production of MG in a concentration- and time- dependent manner. A significant increase in the production of NO, O2.−, and ONOO− was observed in the cells exposed to fructose or MG. Fructose- or MG-induced ONOO− generation was significantly inhibited by MG scavengers and by O2.− or NO inhibitors. The data showed that fructose treatment increased the formation of ONOO− via increased NO and O2.− production in A-10 cells, and this effect was directly mediated by an elevated intracellular concentration of MG. By inhibiting complex III and manganese superoxide dismutase activities, MG induced mitochondrial overproduction of O2.-, and mitochondrial ONOO- further. MG also reduced mitochondrial ATP synthesis, indicating the dysfunction of mitochondria. In addition, MG increased plasma NO levels in patients with T2DM, which reflected the oxidative status in those patients. MG-induced oxidative stress in patients with T2DM significantly enhanced levels of cytokines released from neutrophils. Moreover, the neutrophils from T2DM patients showed a greater proclivity for apoptosis, which was further increased by in vitro MG treatment. Our data demonstrate that MG-induced oxidative damage, particularly ONOO- production, contributes to the pathogenesis of T2DM and its vascular complications.

Canadian statistics indicate that the incidence of obesity is rising, and that the prevalence of type 2 diabetes mellitus (T2DM) within this group is significantly higher than those of a healthy weight. Preliminary evidence has shown that the oil-rich grain, Salvia hispanica L. (Salba), improves glycemic control, suppresses appetite, and affects additional cardiovascular disease (CVD) risk factors. This study followed a randomized, double-blind, placebo-controlled, parallel design in a sub-set population of twenty individuals who were overweight or obese and had T2DM. Participants received supplements of Salba, or an energy- and fibre-matched control, and followed a hypocaloric diet for 24 weeks. Findings of this study reveal that Salba does not significantly affect weight loss, glycemic control or other CVD risk factors. These findings are preliminary and highlight the complexities of weight loss research. Further investigation into the potential health benefits of Salba is currently being carried out.

The pathogenicity of plant pathogen Pseudomonas syringae depends on the type III secretion system which translocates effector proteins into host cells. In response, plants have evolved resistance proteins to detect presence of specific effectors and activate defense responses. The constant host surveillance imposes a strong selective pressure on effector proteins to evolve rapidly in order to evade detection. The P. syringae HopZ1 effector has evolved into three allelic forms as a result of diversifying selection. In this thesis, I aimed to investigate how sequence divergence contributes to the distinct allelic specificities of HopZ1. Mutational analysis of HopZ1a identified three amino acid residues that were potentially involved in dampening host defense responses, and two HopZ1a mutants partially lost the ability to trigger defense responses yet did not lose their virulence functions. These results suggested that distinct host targets could be involved in the defense-eliciting activity and virulence function of HopZ1a.