La mucoviscidose est une maladie génétique caractérisée par une inflammation pulmonairepersistante du poumon résultant d’un recrutement massif de polynucléaires neutrophiles quisécrètent trois protéases à sérine, l’élastase leucocytaire, la protéase 3 et la cathepsine G. Ledéséquilibre de la balance protéases/antiprotéases au site inflammatoire contribue à la protéolyse dutissu pulmonaire et provoque à terme l’insuffisance respiratoire des patients. Les stratégiesthérapeutiques utilisant des inhibiteurs endogènes n’ont pas abouti aux résultats espérés et n’ontciblé que l’élastase soluble. Nous avons étudié l’activité et la régulation des protéases duneutrophile dans les expectorations des patients atteints de mucoviscidose Nous avons démontrél’importance des pièges neutrophiliques extracellulaires (NETs) dans leur séquestration, ce quiexplique leur résistance à l’inhibition et en conséquence leur pouvoir de destruction du tissupulmonaire. Nous avons également démontré qu’un traitement par la DNase facilite l’action desinhibiteurs ce qui ouvre de nouvelles stratégies de thérapie anti-inflammatoire. / No summary available

archives@tulane.edu / Ethanol consumption is known to increase the prevalence and severity of respiratory infection and impaired immunity. The relationships between ethanol exposure, vitamin D levels, the anti-microbial protein cathelicidin/LL-37, other chemokines and cytokines, and their roles in pulmonary infections are explored in this report. Information from experimental model systems that included pulmonary cell culture, mice, and primates were compared to data from humans with alcohol use disorder (AUD) and normal controls. Cathelicidin/LL-37 levels were reduced by ethanol exposure of Nuli-1 airway epithelial cells, which is consistent with the model of enhanced airway sensitivity to infection. The vitamin D receptor positively regulates cathelicidin/LL-37 levels. A reduction in the circulating vitamin D in the Rhesus monkey model following ethanol exposure that may impact cathelicidin/LL-37 expression. Serum samples from ethanol exposed mice, Rhesus monkeys and humans with AUD showed variable expression patterns with respect to cytokines and chemokine, some of which may relate to changes in immunity and infection sensitivity. Protein microarrays revealed altered inflammatory biomarkers in the ethanol-exposed population with AUD. Principle component analysis-derived clustering methodology signaled the presence of systemic inflammation in AUD subjects. The combination of immunological impairment and persistent inflammatory biomarkers are consistent with the known predisposition of individuals with AUD to the development of respiratory infections and acute respiratory distress syndrome. / 1 / Phanuwat Sriyotha

No description available.

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

published_or_final_version / abstract / Pharmacology / Master / Master of Philosophy

L'alcool est la première cause de maladie hépatique en France, et la maladie alcoolique du foie est responsable de près de 7000 décès par an. La surcharge graisseuse est la troisième cause de maladie hépatique, et s'inscrit dans le cadre plus large du syndrome métabolique. La physiopathologie de ces deux types de maladies hépatiques est très similaire. La stéatose, définie par l'accumulation excessive de triglycérides dans le foie, est la première lésion retrouvée chez les patients. La stéatose peut évoluer vers la stéato‐hépatite lorsqu'elle est associée à une inflammation, une mort hépatocytaire, et à l'initiation d'une réponse fibrogénique. La stéato‐hépatite évolue parfois vers la cirrhose, stade ultime avant le carcinome hépatocellulaire. Il n'existe à ce jour aucun traitement efficace de ces maladies hormis le sevrage alcoolique dans le cadre de la maladie alcoolique du foie et un régime associé à de l'exercice dans le cadre de la maladie hépatique d'origine non‐alcoolique. Il est donc urgent d'identifier de nouvelles stratégies thérapeutiques pour lutter contre la progression de ces maladies.L'activation des cellules de Küpffer, les macrophages résidents du foie, joue un rôle déterminant dans le processus inflammatoire qui initie l'atteinte hépatique. Comme tous les macrophages, les cellules de Küpffer peuvent adopter tout un spectre de phénotypes parmi lesquels on distingue deux extrêmes : le phénotype M1 ou pro‐inflammatoire et le phénotype M2 ou anti‐inflammatoire. Les données de la littérature ainsi que celles préalablement obtenues par le laboratoire d'accueil ont établi les effets délétères d'une polarisation pro‐inflammatoire M1 des cellules de Küpffer sur l'évolution de la maladie hépatique d'origine alcoolique ou non alcoolique. Cependant, aucune étude n'avait examiné l'impact d'une polarisation anti‐inflammatoire (M2) des cellules de Küpffer sur ces maladies.L'objectif de ce travail a été d'évaluer si favoriser la polarisation des cellules de Küpffer vers un phénotype M2 anti‐inflammatoire pouvait limiter la progression des maladies hépatiques d'origine alcoolique ou non alcoolique.Afin de mener à bien ce projet, nous avons combiné l'utilisation de modèles murins de maladie hépatique d'origine alcoolique ou non alcoolique, des approches pharmacologiques et des expériences sur cellules isolées. Ces études ont été complétées par des données obtenues sur des biopsies humaines.Ce travail a permis de démontrer que favoriser la polarisation M2 des cellules de Küpffer protège le foie de la stéatose, de la mort hépatocytaire et de l'inflammation. Ces résultats identifient un nouveau mécanisme anti‐inflammatoire déclenché par les cellules de Küpffer polarisées M2 : l'induction sélective de l'apoptose des cellules de Küpffer polarisées M1. Ce travail ouvre de nouvelles perspectives à l'identification de stratégies pour limiter la progression des maladies hépatiques et pourrait également avoir des retombées plus larges dans le cadre d'autres maladies chroniques inflammatoires ciblant d'autres tissus que le foie. / Summary not transmitted

Thesis (Ph. D.)--West Virginia University, 2007. / Title from document title page. Document formatted into pages; contains x, 212 p. : ill. (some col.). Includes abstract. Includes bibliographical references.

Development of chronic inflammation or autoimmunity may be related to deregulated mechanisms orchestrating successful resolution of inflammation, especially apoptosis of inflammatory cells and their subsequent clearance by macrophages (Mφ). Chronically inflamed sites are characterised by an excess of the key pro-inflammatory cytokine tumor necrosis factor-α (TNF-α) and importantly, TNF-α inhibitors, widely used in the clinical setting for the treatment of rheumatoid arthritis (RA), inflammatory bowel disease and psoriasis, significantly delay disease progression. TNF-α therefore may affect processes implicated in resolution of inflammation. Although TNF-α and pro-inflammatory bacterial products such as lipopolysaccharide (LPS) influence rates of inflammatory cell apoptosis, little is known about their effects on Mφ phagocytosis of apoptotic cells (efferocytosis). In this PhD thesis, the effects of several pro-inflammatory agents (i.e., LPS, lipoteichoic acid (LTA), peptidoglycan (PGN) and TNF-α) on efferocytosis by human blood monocytederived Mφ (MDMφ) have been investigated. LPS, LTA and PGN all inhibited MDMφ efferocytosis in a concentration- and time-dependent manner; however, LPS did not inhibit the uptake of immunoglobulin-G (IgG)-opsonized erythrocytes. Moreover, although TNF-α did inhibit efferocytosis, phagocytosis of IgG-opsonized erythrocytes was not inhibited. Furthermore, the LPS effect was attenuated by dimeric soluble human recombinant TNF receptor-1 (sTNFR1/ Fc), indicating a critical role of TNF-α. Concomitant treatments with monomeric soluble human recombinant TNF receptor-1 (sTNF-R1) or the TNF-α Converting Enzyme (TACE) inhibitor, TOPI-0, only partially reversed the inhibitory effect of LPS. Even though TNF-α release takes place within the first few hours following LPS stimulation, the LPS-induced inhibitory effect occurred only if treatment was performed for 96 hours or longer. Analysis of supernatants obtained from LPS-treated MDMφ revealed that there appears to be interplay between concentrations of TNF-α and interleukin-10 (IL-10) and that these cytokines exert opposing actions on efferocytosis. IL-10 per se increased MDMφ efferocytosis and addition of exogenous IL-10 to LPS-treated samples rescued phagocytosis. The latter effect was associated with the IL-10-induced, concentration-dependent inhibition of TNF-α release. Interestingly, when IL-10 was added to TNF-α-treated MDMφ, only slight augmentation of phagocytosis was observed. Furthermore, when IL-10-mediated effects were blocked by concomitant treatment with anti-human IL-10 receptor 1 antibody (anti-IL-10- R1Ab), the LPS inhibitory effect on phagocytosis was much greater and occurred at 24 hours after treatment. The role of IL-10 on efferocytosis was also investigated using IL-10 deficient murine bone marrow-derived Mφ (BMDMφ). IL-10 deficient BMDMφ, when compared to wild-type, were characterised by a much lower ability to phagocytose apoptotic neutrophils and this effect was independent of culture conditions (control samples and LPS or TNF-α treatments). Finally, effects of the synthetic steroid (dexamethasone) and nonsteroidal anti-inflammatory drugs (NSAID) on MDMφ phagocytosis were examined. Dexamethasone, like IL-10, augmented MDMφ efferocytosis, reversed the inhibitory effects of both LPS and TNF-α, and suppressed LPS-induced production of TNF-α. In contrast NSAID did not increase MDMφ efferocytosis per se. However, preliminary data suggest that aspirin blocks the inhibitory effect of TNF-α on phagocytosis. In summary, it has been determined that prolonged treatment with proinflammatory agents such as LPS, LTA and PGN inhibits MDMφ efferocytosis which may potentially postpone the resolution of inflammation in vivo. I have shown that TNF-α is a key mediator in this process and that IL-10 exerts an important regulatory effect on TNF-α production and consequently on efferocytosis. Furthermore, several approaches have been unveiled to successfully reverse LPS-mediated inhibition of efferocytosis by decreasing either TNF-α production or its inhibitory effect with sTNF-RI/Fc, exogenous IL- 10 or dexamethasone. These findings indicate that TNF-α and other agents which influence efferocytosis may have significance in the resolution phase of inflammation. In addition, presented findings provide important mechanistic information into the potential mode of action of anti-TNF-α agents and steroids and may help to explain their clinical success in the treatment of chronic inflammatory diseases.

Sepsis is a life threatening progression of a trauma or pathogen initiated systemic inflammatory response. Current treatment is supportive and depends mainly on antibiotics, fluids, and the careful administration of oxygen therapy. As sepsis progresses, it becomes a dysregulated inflammatory response, characterised by oxidative stress and excessive production of inflammatory cytokines, mitochondrial dysfunction and loss of antioxidant protection. Previous work on cells and animals has shown that novel antioxidants targeted to mitochondria may have a beneficial effect. To induce an inflammatory response and mitochondrial dysfunction, a human umbilical vein endothelial cell (HUVEC) in vitro mixed sepsis model with 0.2 μg/ml lipopolysaccharide (LPS) plus 20 μg/ml peptidoglycan (PepG) was used in the presence of a mitochondrially targeted vitamin E derivative, MitoVit E, and compared to the non-targeted vitamin E forms, Trolox and DL α-tocopherol acetate. Gene expression analysis was performed by quantitative polymerase chain reaction (qPCR) of the Toll-like receptor (TLR) 2 and 4 pathways from cells treated with the antioxidant +/- LPS/PepG for 4 h. Results showed that MitoVit E differentially regulated 11 genes compared to just four genes for the non-targeted forms of vitamin E. MitoVit E, Trolox and vitamin E were able to blunt IL-6 and IL-8 cytokine response in a dose-dependent manner. Inhibition of NFĸB gene and accessory protein expression was different for each antioxidant investigated along with effects on other inflammatory signalling proteins STAT 3 and MyD88. In addition, the antioxidants regulated radical production to similar extents, but had different effects on the reduced glutathione/oxidised glutathione, mitochondrial metabolic activity, mitochondrial membrane potential, oxygen consumption and mitochondrial number. In conclusion, MitoVit E showed encouraging effects in preventing dysregulation of the inflammatory response, maintaining mitochondrial membrane potential and radical production and normal cell function.

Nanomaterials offer an advantage over traditional biomaterials since cells naturally communicate via nanoscale interactions. The extracellular matrix, for example, modulates adhesion and cellular functions via nanoscale features. Thus incorporating nanofeatures into biomaterials may promote tissue regeneration, however in certain forms and doses nanomaterials can also cause harm. A thorough understanding of cell-nanomaterial interactions is therefore necessary to better design functional biomaterials. This thesis focuses on evaluating the effect of nanofeatures on inflammation using two different models: nanoporous alumina and hydroxyapatite nanoparticles (HANPs). The inflammatory response caused by in vitro exposure of macrophages to nanoporous alumina, with pore diameters of 20nm and 200nm, was investigated. In addition in vivo studies were performed by implantation of nanoporous membranes in mice. In both cases the 200nm pore diameter elicited a stronger inflammatory response. Nanoporous alumina with 20, 100 and 200nm pores were loaded with Trolox, a vitamin E analogue, in order to scavenge ROS produced by primary human polymorphonuclear (PMNC) and mononuclear (MNCs) leukocytes. Unloaded alumina membranes stimulated greater ROS production from PMNCs cultured on 20nm versus 100nm pores. This trend reversed when PMNCs were cultured on Trolox loaded membranes since Trolox eluted slower from 20nm than 100nm and 200nm pores. ROS produced from PMNCs was reduced between 8-30% when cultured on Trolox loaded membranes. For MNCs, ROS production was not affected by pore size. However when the alumina was loaded with Trolox ROS production was quenched by 95%. HANPs with distinct morphologies (long rods, sheets, dots, and fibers) were synthesized via hydrothermal and precipitation methods. The HANPs were then exposed to PMNCs, MNCs, and the human dermal fibroblast (hDF) cell line. Changes in cell viability, ROS, morphology, and apoptotic behavior were evaluated. PMNC and hDF viability decreased following exposure to fibers, while the dot particles reduced MNC viability. Fibers stimulated greater ROS production from PMNCs and MNCs, and caused apoptotic behavior in all cell types. Furthermore, they also stimulated greater capsule thickness in vivo, suggesting that nanoparticle morphology can significantly influence acute inflammation. The outcome of this thesis, confirms the importance of understanding how nanofeatures influence inflammation.