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Obesity is recognized as an important risk factor for the development of many chronic diseases such as hypertension, Type 2 diabetes mellitus (T2DM) cardiovascular disease, cancer, renal disease, neurologic dysfunction, metabolic syndrome and asthma (3, 4). Circulating serum adiponectin levels in obese asthmatics have been reported to be low. Therefore, we aimed to investigate the role of adiponectin in a mucus hypersecretion model and hypothesized that adiponectin would decrease IL-13 induced MUC5AC expression from differentiated NHBE cells and that increasing concentrations of IL-13 would cause a decrease in Adipo R1 and Adipo R2 expression. MUC5AC expression with exposure to adiponectin was not significant. However, mRNA expression of Adipo R1 and Adipo R2 was significantly decreased by stimulation of IL-13 for acute (24 hours) and chronic (14 days) exposure. Therefore, the obese state and specifically IL-13 concentration could play a role in Adipo R1 and Adipo R2 expression within NHBE cells.

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<p><strong>Introduction</strong></p> <p>The airway epithelium, which acts as a protective barrier, is impaired in asthmatic patients and may contribute to abnormal airway function. Chronic inflammation, a feature of asthma, is associated with structural changes in the airway epithelium including the transformation of columnar epithelial cells into mucin secreting goblet cells. Human epithelial cells exposed to Interleukin-13 (IL-13) <em>in vitro</em> resulted in goblet cell metaplasia and a significant drop in transepithelial resistance, indicating that barrier function is impaired.</p> <p><strong>Aim</strong></p> <p>We sought to determine whether goblet cell metaplasia alone is sufficient to impair airway epithelial barrier function <em>in vivo</em>.</p> <p><strong>Methods</strong></p> <p>Female BALB/c mice were infected with an adenovirus to overexpress IL-13, a control adenovirus, or no virus. Barrier integrity was assessed via single-photon emission computed tomography (SPECT) imaging by measuring the dispersion of technetium-labeled diethylene triamine pentaacetic acid (^99mTc-DTPA) out of the lungs over time. Lung sections were stained by Periodic acid-Schiff to detect the presence of mucin-containing goblet cells.</p> <p><strong>Results</strong></p> <p>IL-13 exposure resulted in goblet cell metaplasia and associated airway hyperresponsiveness to methacholine. However, there was no significant increase in dispersion of ^99mTc-DTPA over time from the airways in IL-13 overexpressed mice compared to control mice.</p> <p><strong>Conclusion</strong></p> <p>IL-13 induced goblet cell metaplasia did not impair the airway epithelial barrier to ^99mTc-DTPA in our <em>in vivo</em> mouse model. Therefore, we conclude that epithelial dysfunction to DTPA observed in human asthmatics and in animal models of asthma are not due to IL-13 induced goblet cell metaplasia.</p> / Master of Science (MSc)

Mucosal inflammation in conditions ranging from infective acute enteritis or colitis to inflammatory bowel disease (IBD) is accompanied by alteration in enterochromaffin (EC) cell numbers and serotonin (5-hydroxytryptamine; 5-HT) content in the gut. Previously we had shown that CD4+ T cells, via production of T helper (Th)2 cytokines, regulate EC cell biology in the Trichuris muris-infectious colitis model. I further examined the mechanisms of immuno-endocrine interactions in the context of intestinal inflammation. In chapter 3, utilizing human EC cell line and Trichuris muris-mouse model of infectious colitis we identified a critical role of interleukin (IL)-13, a key Th2 cytokine, in increasing EC cell numbers, tryptophan hydroxylase (TPH)1 expression (rate-limiting enzyme of mucosal 5-HT bio-synthesis), and 5-HT production. In chapter 4, we show that IL-13 driven intestinal inflammation is critically dependent on increased 5-HT production using dextran sulfate sodium (DSS) and dinitrobenzene sulphonic acid (DNBS) models of colitis. In DSS-induced colitis, we were the first to identify the increased production of IL-13 and its pathogenic role as IL-13 knockout (IL-13-KO) mice had less severe inflammation compared to wild-type, which was exacerbated following replenishment of 5-HT in IL-13-KO mice. In chapter 5, biopsy examination revealed, higher mucosal IL-13 expression accompanied inflammation in Crohn's disease (CD), which was additionally associated with increased TPH1, 5-HT receptor (5-HTR)3A, 5-HTR7 and decreased 5-HT transporter (5-HTT) expressions. Moreover, CD patients had elevated plasma and platelet-poor plasma 5-HT levels compared to healthy controls (HCs). Furthermore, 5-HTT polymorphism associated genotypes causing inefficiency in 5-HT re-uptake were more common in our patient cohort than HCs. The findings included in this thesis further emphasize the role of immuno-endocrine interactions in intestinal inflammation, which may be a step toward a better diagnosis or management or even a cure for a disease that is of growing concern, and in understanding IBD pathogenesis. / Dissertation / Doctor of Philosophy (PhD) / The gut produces most of the serotonin found in our body, where it regulates many normal functions. A group of special cells, named enterochromaffin cells, produces nearly all of the serotonin in the gut. In diseases of the gut, especially ones that involve inflammation resulting in symptoms like abdominal pain, diarrhea and bleeding, the number of these cells and serotonin concentration are different from that in the normal gut. I found that these changes are controlled by a particular protein produced by immune cells, called interleukin-13, and alteration in serotonin levels, in turn, contributes to the inflammatory process. Our laboratory experiments with cells and animals establish this connection between interleukin-13 and serotonin in gut inflammation. We further confirm this association between interleukin-13 and serotonin in human inflammatory bowel disease. Moreover, we identify a potential genetic cause of these changes in serotonin concentrations which may ultimately result in inflammatory bowel disease.

Inflammatory processes in, or in close vicinity of, the skeleton often lead to loss of bone tissue. Different cytokines have been shown to be involved as stimulators of inflammatory induced osteoclastic bone resorption. During inflammatory processes also the kallikrein-kinin system is activated, leading to production of kinins that can cause pain, vasodilation and increased permeability of vessels. Kinins can also induce bone resorption in vitro. All cytokines and kinins that stimulate bone resorption stimulate in parallell prostaglandin synthesis, and prostaglandins, per se, have also been shown to induce bone resorption. The aim of this project was to increase the knowledge about the mechanisms involved in the interactions between different inflammatory mediators (i.e. kinins, cytokines and prostaglandins) suggested to be involved in the pathogenesis of inflammatory bone resorbing diseases. Human osteoblasts (MG-63) are equipped with both kinin B1 and B2 receptors linked to prostaglandin release and the stimulation of prostaglandin release are likely mediated via separate molecular mechanisms (Paper I). Activation of B1 or B2 receptors causes synergistic stimulation of PGE2 synthesis induced by either interleukin-1b (IL-1b) or tumour necrosis factor-a (TNF-a) (Paper II). The molecular mechanism involves increased expression of cyclooxygenase-2 (COX-2) and results in synergistic potentiation of receptor activator of NF-kB ligand (RANKL) protein expression. The synergistic interaction is dependent on the activation of NF-kB and the mitogen-activated protein kinases (MAPK) p38 and JNK (Paper II). The synergistic increase in RANKL expression might be an explanation why kinins potentiate IL-1b induced bone resorption, a mechanism likely to be important in inflammation induced bone resorption in diseases such as periodontal disease and rheumatoid arthritis. The synergism between kinins and IL-1b or TNF-a might also be dependent on regulation of kinin receptors, since both IL-1b and TNF-a markedly upregulated B1 and B2 receptors, both at the mRNA level and protein level (Paper III). This upregulation is not further potentiated by the kinins, and different kinin receptor agonists do not regulate the receptors for IL-1b or TNF-a, in MG-63 cells. No other cytokines known to stimulate bone resorption regulates the expressions of B1 and B2 receptors. The IL-1b- or TNF-a-induced enhancements of B1 and B2 receptor expressions involve activation of NF-kB and MAPK. The enhancement of kinin receptors may also be an important mechanism in the synergistic interactions between the two pro-inflammatory cytokines and kinins (paper III). IL-4 and IL-13 are two cytokines that have been shown to inhibit bone resorption. We have shown that COX-2 and both B1 and B2 receptors are down-regulated by IL-4 and IL-13, via a ‘signal transducer and activator of transcription6’ (STAT6) dependent pathway, which might be an important regulatory mechanism in inflammation induced bone resorption (paper IV). In conclusion, the mechanisms behind the synergistic potentiation of prostaglandin formation and increased bone resorption caused by co-stimulation with kinins and IL-1b or TNF-a seem to involve both potentiation of COX-2 and subsequently increased levels of RANKL, as well as upregulation of B1 and B2 kinin receptors. Interestingly, IL-4 and IL-13 decreased the expressions of COX-2 and both B1 and B2 receptors. These events might be important in the regulation of inflammation induced bone resorption in diseases such as periodontitis and rheumatoid arthritis.

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Les neurones nocicepteurs jouent un rôle clé dans la défense de l’organisme. Dans le cas des réactions inflammatoires, ils initient des réflexes protecteurs tels que la toux, les vomissements, où les démangeaisons, et participent à la régulation de plusieurs mécanismes physiologiques, notamment la réponse immunitaire. Ils jouent ainsi un rôle prépondérant dans l’inflammation de type 2, souvent associée aux allergies. Mais les mécanismes qui permettent l’activation de ces neurones dans ce contexte sont encore mal connus. Au cours de ce projet de recherche, nous avons exploré la capacité des neurones nocicepteurs à détecter les signaux immunitaires spécifiquement associés à l’asthme. Nous avons ainsi identifié les caractéristiques des nocicepteurs des voies aériennes. Nous avons également démontré leur sensibilité aux allergènes grâce à l’expression du récepteur aux immunoglobulines de type E, FcεR1, ainsi que leur capacité à modifier leur transcriptome en réponse aux cytokines IL-4 et IL-13. Ces travaux soutiennent l’importance de la communication entre systèmes nerveux et immunitaires, et mettent en évidence de nouvelles cibles pour limiter la contribution neuronale aux réactions allergiques. / Nociceptor neurons play a major role in organism defense. In the context of inflammation, they initiate protective reflexes such as cough, vomiting, or itch, and participate in the regulation of various physiological mechanisms, including the immune response. They notably participate in type 2 inflammation, often associated with allergies. But the mechanisms driving the activation of nociceptor neurons in this context are still elusive. During this research project, we investigated the ability of nociceptor neurons to sense immune signals specifically associated with asthma. We identified the characteristics of airway innervating nociceptors. We also demonstrated their sensitivity to allergens through the expression of the Immunoglobulin E receptor FcεR1, as well as their ability to change their transcriptome in response to IL-4 and IL-13. This work supports the importance of bidirectional communication between the nervous and immune systems and unravels new targets to regulate neuronal contribution to inflammation.