Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal.

The aetiology of IBD is unknown, but available evidence suggests that an aberrant immune response towards the commensal microbial flora is responsible for intestinal inflammation in genetically susceptible individuals. Studies from animal models of intestinal inflammation have greatly advanced our understanding of the immunological basis of IBD. However, translation of results from animal research into human studies is essential in order to improve treatment options and patient quality of life. In this thesis we present the successful introduction of translational studies on human tissue in our laboratory. In particular, we evaluated the role of the IL23/IL17 pathway in the human immune response and its role in IBD. IL23-driven inflammation has been primarily linked to its activity on Th-17 cells; however, work from our laboratory has identified a novel population of IL23-responsive ILC, which are responsible for innate colitis in mice. Here we have analyzed the role of IL23-responsive innate cells in IBD. Our results show increased expression of Th-17 signature genes amongst intestinal CD3- cells in patients with IBD. Furthermore, we observed a marked and selective increase in IL17 producing CD56- ILC in the inflamed intestine of patients with CD. ILC may contribute to intestinal inflammation through secretion of cytokines, such as IL17A and IL17F, and recruitment of other inflammatory cells, representing a novel tissue-specific target for the treatment of IBD. In addition, we present here our preliminary data on the characterization of human intestinal and systemic DC populations. In particular, we aimed to evaluate if in the context of the intestinal microenvironment DC develop specific regulatory features, as observed in murine CD103+ DC. We show that human intestinal DC populations exhibit specific regulatory properties, such as expression of genes associated with TGF-β and RA activity. Furthermore, CD103+ DC are present in the human gut and are characterized by tolerogenic markers. Remarkably, patients with IBD have reduced frequencies of intestinal CD103+ DC, which display a more pro-inflammatory phenotype. Alteration in DC subset composition and functional activity may result in a distort balance between immune effector and regulatory responses, promoting the development of intestinal inflammation.

Inflammation appears to play a major role in the pathogenesis of Major Depression, and understanding the relationship between the immune system and mood disorders is a growing area of research. Patients undergoing cytokine therapy frequently develop depressive-like signs which cease upon termination of treatment. In addition, depressed patients often present with high levels of circulating pro-inflammatory cytokines. There are also significant behavioural correlates between inflammation-associated sickness behaviours and Major Depression, suggesting there is a bidirectional relationship between the immune system and central systems regulating behaviour. Both sickness behaviours in animals and cytokine-induced depression in humans can be reversed with antidepressants targeting the serotonergic (5-HT) system. Against this background, this thesis sought to investigate the role of inflammation in the development of certain specific characteristics of depressive-like states in animal models. Specifically, a single systemic endotoxin challenge (LPS) was used to model acute sickness behaviour and to study the behaviour and molecular effects of this challenge on the 5-HT system. These studies indicated that a single systemic challenge changed the function of the 5-HT system, as well as increasing expression levels of a number of 5-HT-related genes, and markers of inflammation in the CNS. A rodent chronic stress paradigm was employed to investigate whether these changes also occur in animals displaying depressive-like signs. Chronically stressed mice were shown to display significant features of rodent depression; decreased sucrose preference and increased forced swim immobility. These changes were also accompanied by increased expression of 5-HT-related genes and inflammatory markers within the CNS. Importantly, many of the molecular changes observed in LPS-induced sickness behaviour were conserved in the model of stress-induced Major Depression. In particular, increased TNF expression within the CNS was highlighted as a feature in both models. Novel anti-inflammatory agents were used to study the role of TNF in the development of sickness behaviours. Anti-TNF therapy, specifically the fusion protein etanercept, ameliorated sickness behaviour induced by LPS. Together, these data demonstrate that inflammation is capable of significantly changing the 5-HT system, and that stress per se is capable of inducing an inflammatory state within the CNS, which support the thesis that sickness and Major Depression are closely related at a molecular level.

Gestational diabetes mellitus (GDM) is a common complication of pregnancy and population health studies have linked it to impaired cognitive performance in the offspring. GDM triggers inflammatory responses, which can critically affect development of neuronal circuitry. We hypothesized that GDM promotes inflammatory responses in the fetus that can disturb fine-tuning of neuronal networks during early development, resulting in lifelong impaired cognitive functions. The cognitive performance of 15 week old offspring exposed to diet induced GDM were assessed. The brain tissue of the 15 week old and neonatal (E20) offspring were analyzed by immunohistochemistry, western blot, and cytokine assay. Cultured microglial responses to elevated glucose and/or fatty acid levels mimicking GDM associated diabetic conditions were analyzed. Our data reveals chronic neuroinflammation in GDM offspring, which combined with deregulation of microglial functions may explain hippocampal CA1 layer neuronal derangement and synaptic degradation that correlates with impaired cognitive performance of GDM offspring. / October 2016

Dissertation submitted in partial compliance with the requirements for the Masters Degree in Technology: Chiropractic, Durban Institute of Technology, 2005 12, xiii, 84 leaves / Plantar Fasciitis (PF) or “painful heel syndrome” is an inflammation of the plantar fascia at its insertion on the medial calcaneal tubercle. Accounting for 7-9% of total sports injuries, this condition is predominantly due to overuse and is notoriously difficult to treat. Traditionally treatment focused on the resolution of the inflammation with the application of such modalities cross frictions / transverse frictions being the modality of choice. With such modalities there are however limitations which include the detection of the appropriate areas in which treatment should be given as well as the treatment depth achieved. The GISTM, however is an advanced form of soft tissue mobilisation that employs the use of specifically designed stainless steel instruments that, when manually brushed over the skin of the affected area, are thought to detect and release scar tissue, adhesions and fascial restrictions. This complementary technique is hypothesized to work in the same manner as cross friction massage, and is thought to achieve quicker and improved outcomes by its detection of the treatment area(s) as well as improving the depth of treatment application. This assertion was however untested. Therefore the purpose of this study was to determine the efficacy of the Graston Technique Instrument-assisted Soft Tissue Mobilisation (GISTM) in the treatment of Plantar Fasciitis in runners. / M

A system of ordinary differential equations is developed to model the effect of fatty acids on chronic intestinal inflammation that is typical of a Crohn’s patient. Several murine studies have shown an anti-inflammatory response when specific polyunsaturated fatty acids are included regularly in the diet. It is believed that the fatty acids serve as a specific ligand that activates the Peroxisome Proliferator Activated Receptor (PPAR) which is located on multiple cell types that are active in the inflammatory response. The binding of the PPAR results in a suppression of the inflammatory pathway. Results of the model indicate a muted inflammatory response when fatty acids are added regularly to the diet in mild to moderate cases of Crohn’s. Results of mathematical analysis show a stable fixed point with decreased inflammatory markers and pathogen levels when fatty acids are added regularly to the diet.

Sterile inflammation is a key component of a variety of diseases including, gout, arthritis, type 1 diabetes, Alzheimer’s disease and multiple sclerosis (MS). Sterile inflammation induces the recruitment of immune cells via chemokines, such as CCL5 and CXCL10. Expression of these chemokines is dependent on IRF-1. Recently the FDA has approved the use of a pro-drug, FTY720 that after phosphorylation becomes a S1P mimetic for the treatment of MS. This report describes two novel and opposing mechanisms of S1P action in sterile inflammation. First, intracellular S1P acts as a cofactor of cIAP2 that inducesIL-1-dependent K63-polyubiquitination of IRF-1, which leads to the recruitment of immune cells to the site of inflammation. Conversely, extracellular S1P provides a feedback loop that inhibits CXCL10 and CCL5 expression through S1PR2 signaling. Accordingly, immune cell infiltration to sites of sterile inflammation is increased in S1PR2-/- production via calcium-dependent, but cAMP- and PKA-independent mechanisms that likely involve c-Fos expression and unconventional PKC activation. Elevated c-Fos could competitively inhibit CCL5 expression directly or indirectly via blocking IFN production. These two novel pathways highlight unexpected aspects of S1P signaling, and provide potential mechanisms that can be exploited for the improvement of therapeutics for the treatment of MS.

Research regarding the development of cardiovascular disease (CVD) is important because CVD is the leading cause of death in the United States (US) and many countries abroad. Risk factors, such as obesity and psychological stress, should be studied in order to understand contributing factors for CVD and the cellular mechanisms which link risk factors with the development of disease. Specifically, the combined influence of multiple risk factors on inflammation is of interest because many individuals have more than one risk factor, which additively increases an individual’s risk for CVD. Obesity is already characterized by disordered inflammation, which suggests that the additional burden of a psychological stressor could elicit a greater inflammatory response and a greater risk for CVD than either stressor alone. The documents included within this thesis include the significance and specific aims of the study in addition to a review of the relevant literature related to the effects of obesity and acute mental stress (AMS) on endocrine and inflammatory markers. Specifically, this study aims to address IL-1β, IL-1Ra, and leptin following an AMS task in non-obese and obese individuals. Additionally, a manuscript is included which evaluates the change in IL-1β, IL-1Ra, and leptin following a 20 minute AMS task. Variables were examined between groups at baseline and at two time points following AMS. Additionally, the relationships among the changes in the markers following AMS were examined. Appendices include expanded methodology and all questionnaires used.

In the presented study, we demonstrate that the interaction of group IVA cytosolic phospholipase A2 and ceramide-1-phosphate is crucial for production of eicosanoid synthesis in inflammation. Inflammation is a critical component of many disease states including anaphylaxis, cancer, cardiovascular disease, rheumatoid arthritis, diabetes and asthma. Eicosanoids are well established mediators of inflammation, and the initial rate limiting step in the production of eicosanoids is the liberation of arachidonic acid (AA) from membrane phospholipids by a phospholipase A2 (PLA2). The major phospholipase involved in this liberation of AA during the inflammatory response is group IVA cytosolic phospholipase A2 (cPLA2α). Previous studies from our laboratory demonstrated that the bioactive sphingolipid, ceramide-1-phosphate (C1P), binds cPLA2α at a three amino acid sequence, which is located in the cationic β-groove of the C2 domain of cPLA2α. In this study we examined the effects of the genetic ablation of ceramide kinase (CERK) on eicosanoid synthesis, as CERK is the only known enzyme to produce C1P in mammalian systems. We utilized primary mouse fibroblasts (MEFs) and macrophages isolated from CERK-/- and +/+ mice. The ceramide-1-phosphate and eicosanoid profiles were investigated, and both ceramide-1-phosphate and eicosanoid levels in CERK-/- MEFs were found to be dysregulated. This study also presents the development of a global eicosanoid method to analyze eicosanoids via LC-ESI-MS/MS. Using this new analysis method, we demonstrated that there are significant differences in eicosanoid levels in ex vivo CERK-/- cells when compared to wild type counterparts, but the effect of the genetic ablation of CERK on eicosanoid synthesis and the serum levels of C1P was not apparent in vivo.

Mast cells respond to a variety of signals, are associated with both increased inflammation and regulation of the immune response, and are able to interact with a variety of hematopoietic and non-hematopoietic cells. The majority of the work that highlights mast cell pleiotropic abilities has been completed in murine models. Though these models have significantly advanced our understanding of what mast cells can do, they cannot inform us as to what mast cells actually do in human beings. The goal of this dissertation is to assess fully mature, primary human mast cell function beyond the well-defined type 1 hypersensitivity function and place mature human mast cells in the context of interactions with other immune cells. The first project addresses the ability of IFNγ, a historically Th1 associated cytokine, to dramatically alter mast cell phenotype. In particular, IFNγ stimulation allows mast cells to act as antigen presenting cells to CD4+ T cells. The second project describes and addresses the T cell suppressive function of myeloid derived suppressor cells in Mastocytosis, a disease of clonal mast cells.