Saudi Cultural Missions Theses & Dissertations
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Item Restricted Exposure to Allergens and Proinflammatory Mediators Modulate Airway Epithelial Cell Innate Responses, Metabolism, and Physiology(University of Alberta, 2024) Alzahrani, Khadija Rashed; Vliagoftis, HarissiosLungs and airways health and protection depend on the integrity of airway epithelium. Insulin is a growth hormone that through activation of several signaling pathways play a central role to maintain cellular growth and activate metabolism. Airway epithelium is in constant exposure to inhaled agents like pathogens, allergens, pollutants, and particular matters. Airborne allergens like cockroach or house dust mite (HDM) contain proteases that can interact with airway epithelium and initiate immune responses through activation of airway epithelial cells and innate immune cells. Airway epithelium cytokines and innate immune cells promote activation of adaptive immune cells like Th2 lymphocytes that produce proinflammatory mediators like interleukin-4 (IL-4), IL-13, and IL-5. T helper 2 (Th2) cytokines amplify the immune responses and stimulate airway epithelial cells to release CCL chemotactic factors/eotaxins that attract eosinophils to the site of inflammation. Airway epithelium exposure to inhaled irritants and excessive inflammatory responses is known to disrupt epithelium integrity and exacerbate inflammatory responses. Th2 inflammation in asthma is associated with epithelial injury, epithelial remodeling, and metabolic alterations. Increased expression of inflammatory mediators tumor necrosis factor (TNF) and IL-6 in metabolically active tissues have been associated with cellular damage and insulin resistance. Inducible AECs local inflammation was shown to trigger systemic inflammation, insulin resistance, and impaired glucose metabolism, and the severity of inflammation correlated with impaired glucose metabolism. Several studies showed that insulin resistance increased the risk of aeroallergen sensitization and developing asthma-like symptoms. In this project we hypothesized that exposure of airway epithelial cells (AECs) to allergens and proinflammatory mediators may modulate Th2 immune cell responses, induce insulin resistance and metabolic shifts. First, we examined the effect of cockroach and house dust mite on IL-13 and IL-4-induced inflammatory responses. AECs were cultured and stimulated with cockroach or house dust mite, IL-13, IL-4, or a combination of an allergen and IL-13 or IL-4. IL-13 and IL-4 induced effects were measured. Then, we tested the effect of cockroach, house dust mite, TNF or IL-6-induced inflammatory responses on physiological epithelial changes and insulin-induced effects. Epithelial proliferation, resistance, energetic phenotype changes, and insulin induced activation of PI3K/Akt were examined. We showed that serine proteases of cockroach extract prevented IL-13-induced expression of eosinophil chemokine CCL26 from AECs. Depletion of CCL26 was previously shown to delay resolution of airway allergic inflammation which may result in prolonged eosinophilia. Prolonged inflammation was linked to altered metabolism and insulin action in several studies. We showed for the first time that house duct mite and TNF modulated insulin effects in AECs. TNF and HDM changed insulin-induced ATP production in AECs. Additionally, TNF reduced insulin-induced Akt phosphorylation, reduced epithelial barrier function and recovery after injury. Our data suggest that cockroach serine proteases and TNF may interfere with the Th2-mediated proinflammatory effects, regulate AECs energetic phenotype, and induce insulin resistance. These inflammatory and metabolic changes exacerbate the pathogenesis of asthma which may alter immune cell responses.5 0Item Restricted Constitutive Release of Purines in Control of Vascular Contractility(Saudi Digital Library, 2023-11-30) Alqarni, Aali; Ralevic, Vera; Dunn, WilliamMost large and medium sized arteries are surrounded by perivascular adipose tissue (PVAT). PVAT was once considered a passive structural component of the vascular wall, not an active regulator of vascular homeostasis. However, PVAT releases several active mediators that have a paracrine effect on vascular tone. Adipocytes express P2X and P2Y receptors, whose activation influences a number of adipocyte functions including the release of adipokines. Adipocytes can also release nucleotides to control adipocyte activity. It is unknown whether this occurs in PVAT and alters the tone of the adjacent vasculature. The constitutive release of purines in blood vessels is relatively unknown. However, it is important to understand vascular control mechanisms to offer new therapeutic approaches for cardiovascular diseases such as hypertension. The main purpose of this study was to determine if nucleotides (ATP/UTP) are released constitutively from PVAT to regulate the vascular tone of porcine mesenteric artery. Further investigation was performed to examine the possibility that P2X, P2Y, and adenosine receptors may contribute to the vasorelaxant effect induced by AR-C118925XX (a P2Y2 receptor antagonist). Additionally, I investigated whether the nucleotides ATP and UTP activate P2Y receptors to release adiponectin and leptin from 3T3-L1 adipocytes. I also examined whether the anti-contractile effect of PVAT in response to ATP and UTP in porcine splenic arteries may be associated with adiponectin release. Isometric tension recordings were performed using mesenteric arteries obtained from large white hybrid pigs with and without PVAT. A direct vasorelaxant effect of suramin (P2 receptor antagonist), AR-C118925XX and MSG228 (P2Y2 receptor antagonists) was found on U46619-preconstricted vascular tone of porcine mesenteric arteries. Apyrase (metabolises nucleotides) abolished the vasorelaxant response of preii contracted mesenteric arteries to AR-C118925XX. Additionally, apyrase caused a dual effect of contraction and relaxation in pre-contracted mesenteric arteries. ARL67156 (ectonucleotidases inhibitor) directly caused an initial small contraction in U46619- pre-constricted mesenteric artery tone. Further experiments demonstrated that the connexin/pannexin channel inhibitors carbenoxolone and probenecid also caused a relaxation of U46619-pre-constricted vascular tone in porcine mesenteric arteries. In this study, evidence ruled out the involvement of PVAT, endothelium and P2X and adenosine receptors in nucleotides release and responses to P2 receptor antagonists, suggesting that nucleotide release and actions involves the vascular smooth muscle. Relaxation of AR-C118925XX was also observed in further vascular beds and species in this study, specifically porcine coronary and splenic arteries, as well as rat mesenteric arteries. Extracellular ATP was measured by luminescence directly from the Krebs solution in porcine mesenteric arteries. It appears that isolated mesenteric arteries are capable of releasing ATP and the presence of PVAT in this experiment increased the amount of ATP released from isolated mesenteric arteries. However, there were no differences in the effects of P2 receptor antagonists in the presence or absence of PVAT on mesenteric artery tone. The present study also identified the role of extracellular nucleotides ATP and UTP in adiponectin release from 3T3-L1 adipocytes via activation of P2Y2 receptors. Adiponectin release was not regulated by constitutive activation of P2Y2 receptors. In isolated arteries, the presence of PVAT in splenic, but not mesenteric, arteries decreased the level of contraction induced by ATP and UTP. The anti-contractile effects of PVAT on porcine splenic arteries in response to ATP and UTP might be mediated by adiponectin and other relaxant factors. Together, these results demonstrated that endogenous ATP/UTP is released constitutively from vascular smooth muscle via connexins and pannexins to act on vasocontractile P2Y2 receptors.34 0